##### 18^th^ North American Barley Researchers Workshop and 4^th^ Canadian Barley Symposium

## Poster Abstracts

[Breeding for malt and feed quality barley in northern Australia
1](#breeding-for-malt-and-feed-quality-barley-in-northern-australia)

[Glen Fox^1,5^, Jan Bowman^3^, Karyn Onley-Watson^1^, Andrew Skerman^1^,
Gary Bloustein^1^, Alison Kelly^1^, Andy Inkerman^1^, David Poulsen^4^
and Robert Henry^2,5^
1](#glen-fox15-jan-bowman3-karyn-onley-watson1-andrew-skerman1-gary-bloustein1-alison-kelly1-andy-inkerman1-david-poulsen4-and-robert-henry25)

[Milling energy and grain hardness in barley
2](#milling-energy-and-grain-hardness-in-barley)

[G. A. Camm and B.G. Rossnagel 2](#g.-a.-camm-and-b.g.-rossnagel)

[Low phytate barley (*Hordeum* *vulgare L.*) development at the Crop
Development Centre, University of Saskatchewan
3](#low-phytate-barley-hordeum-vulgare-l.-development-at-the-crop-development-centre-university-of-saskatchewan)

[B.G. Rossnagel^1^, T. Zatorski^1^, G. Arganosa^1^and V. Raboy^2^
3](#b.g.-rossnagel1-t.-zatorski1-g.-arganosa1and-v.-raboy2)

[Post-anthesis biomass yield and quality of barley cultivars developed
by Field Crop Development Centre
4](#post-anthesis-biomass-yield-and-quality-of-barley-cultivars-developed-by-field-crop-development-centre)

[Nyachiro\* J.M., J.H. Helm and P.E. Juskiw
4](#nyachiro-j.m.-j.h.-helm-and-p.e.-juskiw)

[Process development for quick cooking barley products
5](#process-development-for-quick-cooking-barley-products)

[Hong Qi^1^; Connie Phillips^1^; M. Eliason^2^; Karen Erin^3^ and Feral
Temelli^4^
5](#hong-qi1-connie-phillips1-m.-eliason2-karen-erin3-and-feral-temelli4)

[PCR detection and quantification of *Fusarium* species
6](#pcr-detection-and-quantification-of-fusarium-species)

[Tajinder S. Grewal, Brian G. Rossnagel, Graham J. Scoles
6](#tajinder-s.-grewal-brian-g.-rossnagel-graham-j.-scoles)

[Can we use Australian identified molecular markers for barley net
blotch resistance in western Canadian barley breeding programs?
7](#can-we-use-australian-identified-molecular-markers-for-barley-net-blotch-resistance-in-western-canadian-barley-breeding-programs)

[Tajinder S. Grewal, Brian G. Rossnagel, Graham J. Scoles
7](#tajinder-s.-grewal-brian-g.-rossnagel-graham-j.-scoles-1)

[Selection for improved scald resistance in the Crop Development Centre
barley improvement program
8](#selection-for-improved-scald-resistance-in-the-crop-development-centre-barley-improvement-program)

[B.G. Rossnagel^1^, D. Voth^1^, T. Zatorski^1^, D.D. Orr^2^ and T.K.
Turkington^2^
8](#b.g.-rossnagel1-d.-voth1-t.-zatorski1-d.d.-orr2-and-t.k.-turkington2)

[Selection for improved FHB tolerance in the Crop Development Centre
barley improvement program
9](#selection-for-improved-fhb-tolerance-in-the-crop-development-centre-barley-improvement-program)

[B. Rossnagel^1^, D. Voth^1^, T. Zatorski^1^, J. Tucker^2^, W. Legge^2^,
and M. Savard^3^
9](#b.-rossnagel1-d.-voth1-t.-zatorski1-j.-tucker2-w.-legge2-and-m.-savard3)

[An AFLP derived tightly linked marker for true loose smut resistance
(*Un*8)
10](#an-aflp-derived-tightly-linked-marker-for-true-loose-smut-resistance-un8)

[Peter Eckstein, Donna Hay, Brian Rossnagel, and Graham Scoles
10](#peter-eckstein-donna-hay-brian-rossnagel-and-graham-scoles)

[Cytological karyotyping of *Pyrenophora teres*
11](#cytological-karyotyping-of-pyrenophora-teres)

[A.D. Beattie, G.J. Scoles, B.G. Rossnagel
11](#a.d.-beattie-g.j.-scoles-b.g.-rossnagel)

[A molecular linkage map of *Pyrenophora teres*
12](#a-molecular-linkage-map-of-pyrenophora-teres)

[A.D. Beattie, G.J. Scoles, B.G. Rossnagel
12](#a.d.-beattie-g.j.-scoles-b.g.-rossnagel-1)

[The barley stem rust resistance gene *Rpg5* encodes NBS-LRR and protein
kinase domains in a single gene
13](#the-barley-stem-rust-resistance-gene-rpg5-encodes-nbs-lrr-and-protein-kinase-domains-in-a-single-gene)

[R. Brueggeman^1^, T. Drader^1^, A. Druka^2^, T. Cavileer^3^, B.
Steffenson^4^, J. Nirmala^1^, H. Bennypaul^1^, K. Gill^1^ and A.
Kleinhofs^1,5^
13](#r.-brueggeman1-t.-drader1-a.-druka2-t.-cavileer3-b.-steffenson4-j.-nirmala1-h.-bennypaul1-k.-gill1-and-a.-kleinhofs15)

[*In vitro* selection for pre-screening barley for resistance to
*Fusarium* head blight
14](#in-vitro-selection-for-pre-screening-barley-for-resistance-to-fusarium-head-blight)

[Kumar, K. (1), Xi, K. (1), Helm, J.H. (2), Turkington, T.K. (3), and
Jennifer Zantinge (1)
14](#kumar-k.-1-xi-k.-1-helm-j.h.-2-turkington-t.k.-3-and-jennifer-zantinge-1)

[Diversification strategies for barley disease management in Alberta
15](#diversification-strategies-for-barley-disease-management-in-alberta)

[Turkington, T.K. (1), Xi, K. (2), Clayton, G.W. (1), Harker, K.N. (1),
O'Donovan, J.G. (1), and Lupwayi, N. (1)
15](#turkington-t.k.-1-xi-k.-2-clayton-g.w.-1-harker-k.n.-1-odonovan-j.g.-1-and-lupwayi-n.-1)

[Resistance of western Canadian barley genotypes to scald in Alberta
16](#resistance-of-western-canadian-barley-genotypes-to-scald-in-alberta)

[K. Xi (1), T.K. Turkington (2) and C. Bos (1)
16](#k.-xi-1-t.k.-turkington-2-and-c.-bos-1)

[Mapping and molecular marker development of scald resistance in 'Seebe'
barley
17](#mapping-and-molecular-marker-development-of-scald-resistance-in-seebe-barley)

[Zantinge, J.L.\*, J.H. Helm, Z. Hartman, J.B. Russell, and K. Xi
17](#zantinge-j.l.-j.h.-helm-z.-hartman-j.b.-russell-and-k.-xi)

[Breeding for multiple disease and multiple gene resistance in barley
18](#breeding-for-multiple-disease-and-multiple-gene-resistance-in-barley)

[James H. Helm^1^, H. Vivar^2^, F. Capettini^2^, K. Xi^1^, P. Juskiw^1^,
and J. Zantinge^1^
18](#james-h.-helm1-h.-vivar2-f.-capettini2-k.-xi1-p.-juskiw1-and-j.-zantinge1)

[Variation in virulence among net blotch isolates infecting barley
19](#variation-in-virulence-among-net-blotch-isolates-infecting-barley)

[Linnea G Skoglund 19](#linnea-g-skoglund)

[Assessment of artificial inoculation methods and deoxynivalenol levels
in barley lines representing various candidate sources of resistance to
*Fusarium* head blight
20](#assessment-of-artificial-inoculation-methods-and-deoxynivalenol-levels-in-barley-lines-representing-various-candidate-sources-of-resistance-to-fusarium-head-blight)

[Geddes, J. (1), Eudes, F. (1), Legge, B. (2), Tucker, J. (2)
20](#geddes-j.-1-eudes-f.-1-legge-b.-2-tucker-j.-2)

[Reactions of barley lines to leaf rust, caused by *Puccinia hordei*
21](#reactions-of-barley-lines-to-leaf-rust-caused-by-puccinia-hordei)

[Y. Sun (1), J. D. Franckowiak (2), and, S. M. Neate (1)
21](#y.-sun-1-j.-d.-franckowiak-2-and-s.-m.-neate-1)

[The differences in fermentable carbohydrates of major Canadian malting
barley varieties and their effects on fermentation
22](#the-differences-in-fermentable-carbohydrates-of-major-canadian-malting-barley-varieties-and-their-effects-on-fermentation)

[Yueshu Li^1^, Rob McCraig^1^, Ken Sawatzky^1^, Aleks Egi^1^ and Michael
Edney^2^
22](#yueshu-li1-rob-mccraig1-ken-sawatzky1-aleks-egi1-and-michael-edney2)

[NanoMash: A novel procedure for research mashing of limited-quantity
barley malts
22](#nanomash-a-novel-procedure-for-research-mashing-of-limited-quantity-barley-malts)

[Laurie A. Marinac and Mark R. Schmitt\*
22](#laurie-a.-marinac-and-mark-r.-schmitt)

[Comparison of hull peeling resistance of barley and malt in western
Canadian two-row barley lines
23](#comparison-of-hull-peeling-resistance-of-barley-and-malt-in-western-canadian-two-row-barley-lines)

[W.G. Legge^1^, J.S. Noll^2^, and B.G. Rossnagel^3^
23](#w.g.-legge1-j.s.-noll2-and-b.g.-rossnagel3)

[Elimination of barley colour defects in Australia
24](#elimination-of-barley-colour-defects-in-australia)

[Glen Fox^1^, Maria Sulman^1^ and Kevin Young^2^
24](#glen-fox1-maria-sulman1-and-kevin-young2)

[Characterization of barley tissue-ubiquitous beta-amylase2
24](#characterization-of-barley-tissue-ubiquitous-beta-amylase2)

[Suzanne E. Clark^1^, Patrick M. Hayes^2^, and Cynthia A. Henson^1,3^
24](#suzanne-e.-clark1-patrick-m.-hayes2-and-cynthia-a.-henson13)

[Barley seed osmolyte concentration as an indicator of preharvest
sprouting
25](#barley-seed-osmolyte-concentration-as-an-indicator-of-preharvest-sprouting)

[Cynthia A. Henson^1,2^, Stanley H. Duke^2^, Paul Schwarz^3^, Rich
Horsley^3^, and Charles Karpelenia^1^
25](#cynthia-a.-henson12-stanley-h.-duke2-paul-schwarz3-rich-horsley3-and-charles-karpelenia1)

[Osmolyte concentration as an indicator of malt quality
25](#osmolyte-concentration-as-an-indicator-of-malt-quality)

[Cynthia A. Henson^1,2^, Stanley H. Duke^2^, and Charles Karpelenia^1^
25](#cynthia-a.-henson12-stanley-h.-duke2-and-charles-karpelenia1)

[Relationships among malt fermentability and malt quality parameters
under the influence of barley β-amylase heat stable allele
26](#relationships-among-malt-fermentability-and-malt-quality-parameters-under-the-influence-of-barley-β-amylase-heat-stable-allele)

[Blanca Gómez\*^1^, Héctor Acevedo^2^, and Ana Clara López^3^
26](#blanca-gómez1-héctor-acevedo2-and-ana-clara-lópez3)

[Russian wheat aphid resistant barley -- cultivar and germplasm release
27](#russian-wheat-aphid-resistant-barley-cultivar-and-germplasm-release)

[D.W. Mornhinweg^1^, P.P. Bregitzer^2^, D.A. Obert^2^, F.B. Peairs^3^,
D. Baltensperger^4^ and R. Hammon^3^
27](#d.w.-mornhinweg1-p.p.-bregitzer2-d.a.-obert2-f.b.-peairs3-d.-baltensperger4-and-r.-hammon3)

[BARMS: A new relational database for barley breeding programs
28](#barms-a-new-relational-database-for-barley-breeding-programs)

[D. B. Cooper and Bruce Westlund 28](#d.-b.-cooper-and-bruce-westlund)

[Mapping and molecular marker development of seed dormancy in a barley
population derived from 'Samson' barley
29](#mapping-and-molecular-marker-development-of-seed-dormancy-in-a-barley-population-derived-from-samson-barley)

[J.L. Zantinge\*, J.M. Nyachiro, S. Chisholm, J.H. Helm, P.E. Juskiw and
D.F. Salmon
29](#j.l.-zantinge-j.m.-nyachiro-s.-chisholm-j.h.-helm-p.e.-juskiw-and-d.f.-salmon)

[Genotypic variations in preharvest sprouting resistance and seed
dormancy in barley
30](#genotypic-variations-in-preharvest-sprouting-resistance-and-seed-dormancy-in-barley)

[Nyachiro\* J.M., J.L. Zantinge, J.H. Helm, P.E. Juskiw and D.F. Salmon
30](#nyachiro-j.m.-j.l.-zantinge-j.h.-helm-p.e.-juskiw-and-d.f.-salmon)

[Using growing degree days to estimate maturity in small grain cereals
31](#number-range-chapterusing-growing-degree-days-to-estimate-maturity-in-small-grain-cereals)

[Juskiw, P., Helm, J., Salmon, D., and Nyachiro, J.
31](#juskiw-p.-helm-j.-salmon-d.-and-nyachiro-j.)

[Twelve years of barley-based rotations
31](#twelve-years-of-barley-based-rotations)

[Juskiw, P., and Westling, D. 31](#juskiw-p.-and-westling-d.)

[Development of winter hulless barley varieties as a high value crop
32](#development-of-winter-hulless-barley-varieties-as-a-high-value-crop)

[W.S. Brooks\*, C.A. Griffey and M.E. Vaughn
32](#w.s.-brooks-c.a.-griffey-and-m.e.-vaughn)

[Multiple dominant and recessive marker stock development
33](#multiple-dominant-and-recessive-marker-stock-development)

[Robert I. Wolfe 33](#robert-i.-wolfe)

[Genetic male sterile and xenia assisted reciprocal recurrent selection
34](#genetic-male-sterile-and-xenia-assisted-reciprocal-recurrent-selection)

[Robert I. Wolfe 34](#robert-i.-wolfe-1)

[Isoyield analysis of barley cultivar trials in the Canadian Prairies
35](#isoyield-analysis-of-barley-cultivar-trials-in-the-canadian-prairies)

[Rong-Cai Yang^1,2\*^, Daniel Stanton^2,3^, Stanford F. Blade^4^, James
Helm^5^ and Dean Spaner^2^
35](#rong-cai-yang12-daniel-stanton23-stanford-f.-blade4-james-helm5-and-dean-spaner2)

[Genetic analysis of preharvest sprouting in barley
36](#genetic-analysis-of-preharvest-sprouting-in-barley)

[S.E. Ullrich\*, J.A. Clancy, H. Lee, F. Han, K. Matsui, I.A. del Blanco
36](#s.e.-ullrich-j.a.-clancy-h.-lee-f.-han-k.-matsui-i.a.-del-blanco)

[Effects of ethylene in barley (*Hordeum vulgare* L.) tissue culture
regeneration
37](#effects-of-ethylene-in-barley-hordeum-vulgare-l.-tissue-culture-regeneration)

[Jha, Ajay K.^1^, Lynn S. Dahleen^2^ and Jeff C. Suttle^2^
37](#jha-ajay-k.1-lynn-s.-dahleen2-and-jeff-c.-suttle2)

[Validation of select diastatic power QTL in elite Western U. S.
six-rowed spring barley germplasms
38](#validation-of-select-diastatic-power-qtl-in-elite-western-u.-s.-six-rowed-spring-barley-germplasms)

[D. Hoffman, A. Hang, and D. Obert 38](#d.-hoffman-a.-hang-and-d.-obert)

[The barley stem rust resistance gene product RPG1 is specifically
degraded upon infection with the stem rust fungus *Puccinia graminis* f.
sp. *tritici* pathotype MCC
39](#the-barley-stem-rust-resistance-gene-product-rpg1-is-specifically-degraded-upon-infection-with-the-stem-rust-fungus-puccinia-graminis-f.-sp.-tritici-pathotype-mcc)

[J. Nirmala^1^, B. Steffenson^2^ and A. Kleinhofs^1&\ 3\*^
39](#j.-nirmala1-b.-steffenson2-and-a.-kleinhofs1-3)

[Saturation mapping of barley chromosome 2H *Fusarium* head blight
resistance QTL
40](#saturation-mapping-of-barley-chromosome-2h-fusarium-head-blight-resistance-qtl)

[Christina Maier^1^, Deric Schmierer^1^, Thomas Drader^1^, Richard
Horsley^2^, and Andris Kleinhofs^1,3^
40](#christina-maier1-deric-schmierer1-thomas-drader1-richard-horsley2-and-andris-kleinhofs13)

[Map-based cloning efforts of the barley spot blotch resistance gene
*Rcs5*
41](#map-based-cloning-efforts-of-the-barley-spot-blotch-resistance-gene-rcs5)

[Thomas B Drader^1^, Kara A Johnson^2^, Robert S Brueggeman^1^, Hye Ran
Kim^3^, Dave Kudrna^3^, Rod Wing^3^, Brian Steffeson^4^, and Andris
Kleinhofs^1,\ 5^
41](#thomas-b-drader1-kara-a-johnson2-robert-s-brueggeman1-hye-ran-kim3-dave-kudrna3-rod-wing3-brian-steffeson4-and-andris-kleinhofs1-5)

[A gene tagging system for *Hordeum vulgare*
42](#a-gene-tagging-system-for-hordeum-vulgare)

[François Eudes^1\*^, André Laroche^1^, Michele Frick^1^, Jennifer
Geddes^1^ and Laurian Robert^2^
42](#françois-eudes1-andré-laroche1-michele-frick1-jennifer-geddes1-and-laurian-robert2)

[Unraveling the mysteries of germination using SAGE (Serial Analysis of
Gene Expression)
43](#unraveling-the-mysteries-of-germination-using-sage-serial-analysis-of-gene-expression)

[Toni Pacey-Miller, Jessica White, Allison Crawford, Peter Bundock,
Giovanni Cordeiro, Daniel Barbary and Robert Henry
43](#toni-pacey-miller-jessica-white-allison-crawford-peter-bundock-giovanni-cordeiro-daniel-barbary-and-robert-henry)

## Breeding for malt and feed quality barley in northern Australia

### Glen Fox^1,5^, Jan Bowman^3^, Karyn Onley-Watson^1^, Andrew Skerman^1^, Gary Bloustein^1^, Alison Kelly^1^, Andy Inkerman^1^, David Poulsen^4^ and Robert Henry^2,5^

#### ^1^ Department of Primary Industries& Fisheries, Toowoomba, Queensland, Australia ^2^ Grain Foods Cooperative Research Centre, Lismore NSW, Australia ^3^ Montana State University, Bozeman, Montana, USA ^4^ Department of Primary Industries & Fisheries, Warwick, Queensland, Australia ^5^ Southern Cross University, Lismore NSW, Australia.

Most countries that produce barley classify their varieties as either
malt or feed with the feed class consisting of varieties that are not
biochemically suited for malting. However, these varieties have probably
not been tested for any animal feed value. In a number of countries,
including Australia, more barley is used annually for feeding animals
than used in beer production. Under Australian feedlot conditions,
anecdotal data had suggested that malt varieties were best for feeding
cattle but little data was available to support this generalisation. We
have undertaken a study comparing over 30 Australian varieties and
breeding lines to ascertain some scientific basis to this theory.
Genotypes from two sites and two years replicated trials were evaluated
for malt and feed analysis. Results indicated that the levels of resting
grain components were similar for each end-use. There was no apparent
difference in total starch content between malt and feed. However, there
were differences for the *in sacco Dry Matter Digestibility* with the
good feed and malt genotypes having low levels. While there was no
strong relationship for particle size (hardness) between malt and feed
quality there was a relationship within a genotype with feed type being
slightly harder. This relationship was independent of protein content.
The most significant area of difference is the need for malt varieties
to produce moderate to high levels of enzymes to breakdown endosperm
components during malting and mashing. Varieties that performed
especially well in both end-uses, ie good malt quality and improved
animal performance, were current malting varieties. The biochemical
results to date demonstrate that breeding programs could effectively
select for improved malt and feed quality in breeding lines by focusing
on malt quality and selecting lines with high level of enzymes.

Corresponding author: glen.fox@dpiq.ld.gov.au

## Milling energy and grain hardness in barley

### G. A. Camm and B.G. Rossnagel

#### Plant Sciences Department/Crop Development Centre, University of Saskatchewan, 51 Campus Drive, Saskatoon, Saskatchewan S7N 5A8

Grain hardness is a product of the complex interaction between
compositional and structural endosperm components, including starch,
protein and beta-glucan. Hardness may contribute significantly to barley
quality. Grain hardness can be evaluated by measuring the energy
required to mill (milling energy) or crush (hardness) the grain, with
harder grain requiring more force. Our research examines the
relationship between milling energy and hardness of several feed and
malting barley genotypes grown at multiple locations and the influence
of protein and moisture on grain hardness.

Seven feed, one malting variety and one malting barley breeding line
were grown in field trials at six Western Canadian sites during 2003 and
2004 and evaluated for milling energy, hardness, moisture and protein
content. Milling energy was determined using the 'Comparamill' at the
Scottish Crop Research Institute (Scotland). Hardness and moisture were
determined using the Perten Single Kernel Characterization System
(SKCS). Grain protein was estimated using Near Infrared Transmittance
(NIT).

Analysis of variance showed significant differences between genotypes
and sites for all measured traits (P = \<0.001) with no variety by site
interaction (P = \>0.99). Milling energy of genotypes ranged from 617 to
736 joules (SE = 5.9). McLeod and CDC Dolly required significantly more
energy to mill, followed by Valier, Newdale, Xena and CDC Helgason. CDC
Bold, TR253 and CDC Trey required the least energy to mill, indicating a
softer endosperm. Milling energy ranged from 625 to 709 joules across
sites. SKCS hardness of genotypes ranged from 38.5 to 56.6 (SE = 0.77).
McLeod was hardest, followed by Valier, Xena and CDC Dolly. CDC Trey,
Newdale, TR253, CDC Helgason, and CDC Bold followed with CDC Bold being
softest. SKCS hardness ranged from 40.4 to 55.8 across sites. Protein
concentration of genotypes ranged from 10.8 to 12.0% (SE = 0.16).
McLeod, CDC Dolly and Newdale were highest followed by Valier, CDC
Helgason, TR253, CDC Bold, Xena and CDC Trey. Protein concentration
ranged from 8.8% to 13.3% across sites. Moisture of genotypes ranged
from 10.1% to 10.5% (SE = 0.06), with larger differences between sites
(7.4% to 13.1%). Milling energy was correlated (n = 9) with SKCS
hardness (r = 0.81, P = \<0.008) and protein concentration (r = 0.79, P
= \<0.01). No significant correlation was detected between milling
energy and moisture (P = 0.90), SKCS hardness and moisture (P = 0.89) or
SKCS hardness and protein concentration (P = 0.20).

## Low phytate barley (*Hordeum* *vulgare L.*) development at the Crop Development Centre, University of Saskatchewan

### B.G. Rossnagel^1^, T. Zatorski^1^, G. Arganosa^1^and V. Raboy^2^

#### ^1^Department of Plant Sciences/Crop Development Centre, University of Saskatchewan, Saskatoon, SK, CANADA, S7N 5A8 ^2^U.S.D.A. Agricultural Research Service, National Small Grains Research Facility, 1691 So. 2700 W., Aberdeen, ID 83210, USA

Phytate, a complex of phytic acid (myo-inositol
1,2,3,4,5,6-hexakisphosphate) and other minerals, is the primary form of
phosphorous in barley grain. Monogastric animals do not effectively
digest phytate because they do not produce the phytase enzyme. Diets
must be supplemented with inorganic phosphorous (P) or a microbial
phytase to meet minimum nutritional requirements. Consequently, excreted
phytate generates high levels of P in effluent resulting in possible
environmental pollution or eutriphication of waterways. Low phytic acid
mutants (with corresponding increases in free and available phosphorous)
have been developed in Harrington barley by Dr. V. Raboy, U.S.D.A.
Hvlpa1-1 has 50% less phytate and M635 and M955 have 75% and 95% less,
respectively. Initial hybridizations of the low phytate genotypes were
made in 1998 at the Crop Development Centre (CDC) to adapted hulless
parents in the combinations: Hvlpa1-1/CDC McGwire and M635/CDC Freedom.
Based on the uniformity of the original mutants from an observation
trial in 1999 the initial crosses were subjected to a rapid backcross
breeding strategy with CDC McGwire and CDC Freedom as recurrent parents.
Four backcrosses were made for each hybrid combination between 1999 and
2000 with each F~1~ being screened for phytate to retroactively identify
the low phytate F~1~ plants for the correct backcross in the greenhouse
at the University of Saskatchewan. BC~4~F~1~ generations were grown as
bulk populations in 2000/01 in New Zealand winter nurseries and the
subsequent F~2~ populations were grown as space planted bulks at
Saskatoon, SK in 2001. The BC~4~F~3~ and BC~4~F~4~ generations were
advanced using a modified single seed descent procedure in the
greenhouses at the University of Saskatchewan during the 2001/02 winter.
BC~4~F~5~ lines were grown in the field at Saskatoon as F~5~ hill plots
in 2002. Each hill plot was derived from an individual F~4~ head.
Selected hills were tested for phytate. Seed from selected low phytate
F~5~ hill plots was bulked and increased in 2002/03 winter nurseries in
New Zealand. Selections were tested in CDC yield trials in 2003, two of
which, SR03013 (50% phytate reduction) and SR03044 (75% phytate
reduction), were advanced to the Western Canadian Hulless Barley
Cooperative (WCHBCoop) yield trial during 2004 as HB378 and HB379,
respectively. HB379 has been advanced for final year testing in the 2005
WCHBCoop and will be put forward for support for variety registration in
2006. Growing 2^nd^ generation Breeder Hills in our 2004/05 New Zealand
contra-season nursery has allowed for rapid production of Breeder Seed
of HB379 in 2005 in anticipation of variety registration. Using this
rapid breeding technique means we have moved from 1^st^ cross in 1998 to
a released variety in 2006, a period of less than eight years.

E-mail: Brian.Rossnagel@usask.ca

## Post-anthesis biomass yield and quality of barley cultivars developed by Field Crop Development Centre

### Nyachiro\* J.M., J.H. Helm and P.E. Juskiw

#### Field Crop Development Centre, Alberta Agriculture, Food and Rural Development, 5030 -- 50 St., Lacombe, AB T4L 1W8 Web Site: http://www1.agric.gov.ab.ca/app21/rtw/selsubj.jsp

\*Corresponding author: joseph.nyachiro@gov.ab.ca

Over 2.5 million tonnes of barley silage is produced each year in
Alberta to support the livestock industry. Barley is a vigorous, early
maturing crop that makes high quality silage and is also a preferred
feed grain for Alberta producers. The objective of this study was to
determine post-anthesis (PA) biomass yield and quality of barley
varieties and advanced breeding lines developed at the Field Crop
Development Centre (FCDC), Lacombe. Tests were grown at Lacombe from
1998 to 2004, excluding data for the drought year 2002. The varieties
were grown in replicated field trials. At about soft-dough growth stage
(post-anthesis) the plots were harvested and wet weights determined.
Samples were analyzed to determine quality and percent moisture so dry
weight could be calculated. Sub-samples of biomass were analyzed for
percent protein, acid detergent fibre (ADF%), neutral detergent fibre
(NDF%) and relative feed value (RFV) was calculated. Overall, there were
significant variations of 5 to 20 tonnes/ha of PA biomass yield among
barley varieties. On average there were no significant differences
between the 2-rowed, 6-rowed or hulless barley classes. Biomass protein
for all varieties ranged between 8 and 15%. The 6-rowed and hulless
barley classes tended to have slightly wider range of protein values
compared with the 2-rowed. The ADF ranged between 20 and 40%. The
overall NDF ranged between 30 and 65%, although the 2-rowed barleys
showed a relatively narrower NDF range of between 39 and 59%. The
overall RFV varied between 85 and 200. The 2-rowed barleys showed
narrower RFV values varying between 100 and 160 compared with either
6-rowed or hulless barleys. The ADF was positively correlated (*r* =
0.85) with NDF, and grain yield was positively correlated (*r* = 0.75)
with biomass yield. The biomass yields, protein % and grain yield showed
no correlation to ADF, NDF or RFV. These results suggest that it is
possible to breed barley for high post-anthesis biomass yields and
quality.

## Process development for quick cooking barley products

### Hong Qi^1^; Connie Phillips^1^; M. Eliason^2^; Karen Erin^3^ and Feral Temelli^4^

#### ^1^ Centre for Agri-Industrial Technology, Processing Division, Alberta Agriculture, Food & Rural Development (AAFRD) ^2^ Agricultural Engineering Branch, Technical Services, AAFRD ^3^ Processing Division, AAFRD ^4^ University of Alberta

Barley is among the most ancient of the cereal crops. Canada is the
world's third largest barley producer with an average annual production
of 12 million tonnes. Alberta produces approximately one half of the
total Canadian production. A large percentage of the production is used
as feed for cattle, swine and poultry, while the second largest usage is
in the malting industry. A limited amount of barley is used for human
food. The objective of this study was to develop processes for
quick-cooking barley products to increase food barley consumption.

Alberta grown Falcon and AC Metcalfe cultivars were used for the study.
The effects of variety, pearling rate and pre-treatment on moisture
uptake were studied. Moisture uptake was used to evaluate the effect of
each pre-treatment. Two processes were developed to produce
quick-cooking barley. The quick-cooking barley products reduced cooking
time from 45 minutes to 15 minutes for 35% pearled barley and from 60
minutes to 18 minutes for 5% pearled barley. Quick-cooking barley can be
cooked by boiling in at least twice the volume of water for 15 to 18
minutes followed by a 5-minute stand. Quick-cooking barley products have
a cooked texture, as measured by an Ottawa extrusion method, with an
average force of 450 N, an average bulk density of 530 kg/m^3^ and an
appearance similar to long-cooking barley (cooking 35% pearled barley
for 45 minutes and 5% pearled barley for 1 hour).

There was a slight decrease in the beta-glucan content with the
treatments except with the 35% pearled AC Metcalfe, where the
beta-glucan level increased. The 5% pearled barley had much higher
insoluble dietary fiber content than that of the 35% pearled barley
samples. There was a slight increase in the soluble fibre content with
the pressure treatment compared with the untreated barley samples.

There were no significant differences between the samples for overall
acceptability or flavour, but appearance and texture were significantly
different. The steamed barley samples scored significantly higher for
appearance and colour than the pressure-cooked samples. The
pressure-cooked barley scored significantly higher than the steam-cooked
samples for texture, bite and stickiness/looseness.

The successful development of a quick-cooking barley process provides an
excellent commercialization opportunity for processors to produce a
human consumption barley product, which could be conveniently
incorporated into our daily diet.

Hong.qi@gov.ab.ca

## PCR detection and quantification of *Fusarium* species

### Tajinder S. Grewal, Brian G. Rossnagel, Graham J. Scoles

#### Crop Development Centre/Department of Plant Sciences, University of Saskatchewan, 51 Campus Drive, Saskatoon, SK S7N 5A8 Canada

Fusarium head blight (FHB) is presently the most significant disease of
barley in Canada and many areas of the world. FHB is caused by many
*Fusarium* species and it is very difficult to correlate visual symptoms
with deoxynivalenol (DON) concentration as symptomless kernels may carry
the pathogen. Conversely many blighted kernels may not be contaminated
with DON. Symptoms may be caused by non-trichothecene producing
*Fusarium* species. Identification of *Fusarium* species based on
morphological characteristics by growing on selective media is
cumbersome and requires considerable expertise and experience. In
addition DON estimation is slow, labour intensive and expensive and
demand for this service outstrips supply. PCR-based assays to quantify
fungal DNA in infected plant tissue could indirectly estimate DON
levels. PCR assays have been standardized to detect *Fusarium* species
with species--specific primers (reported by Parry and Nicholson 1996,
Schilling et al. 1996, Nicholson et al. 1998, Yoder and Christianson
1998, Aoki and O\'Donnell 1999, Williams et al. 2002) using 22 isolates
of seven different *Fusarium* species. The protocol to detect *F.
graminearum* associated with FHB symptoms produced under artificial and
natural infections has been standardized. Primer-pair Fg16N amplified
the desired band (280 bp) in all infected barley samples from a
greenhouse experiment and no band was amplified from uninoculated and
uninfected samples. The vast majority of samples from artificially and
naturally infected samples from barley fields showed the desired band.
Protocol to quantify *F. graminearum* DNA using a competitor DNA
template (obtained from Dr. P. Nicholson, John Innes Centre, Norwich,
UK) has been standardized. Fungal concentration ranging from 1 pg to 100
ng with a constant competitor DNA template concentration (1fg or 3fg/μl)
was evaluated and fungal DNA concentration as low as 100 pg (0.1ng) was
detected. Work is in progress to quantify fungal DNA from FHB infected
barley.

##### References

Aoki, T. and O\'Donnell. 1999. *Mycologia*. 91:597-609.

Nicholson P. et al. 1998. *Physiological and Molecular Plant Pathology*.
53:17-37.

Parry D. and Nicholson P. 1996. *Plant Pathology*. 45:383-391.

Schilling A.G. et al. 1996. *Phytopathology*. 86:515-522.

Williams K.J. et al. 2002. *Australian Plant Pathology*. 31: 119-127.

Yoder, W. and Christianson, L. 1998. *Fungal Genetics and Biology*. 23:
68-80.

## Can we use Australian identified molecular markers for barley net blotch resistance in western Canadian barley breeding programs?

### Tajinder S. Grewal, Brian G. Rossnagel, Graham J. Scoles

#### Crop Development Centre/Department of Plant Sciences, University of Saskatchewan, 51 Campus Drive, Saskatoon, SK S7N 5A8 Canada

Net blotch of barley caused by Pyrenophora teres Drechs. is an important
disease in western Canada (Tekauz 1990) and elsewhere (Steffenson 1997).
Two types of leaf symptoms are associated with the net blotch disease:
the net form (NFNB), caused by P. teres f. teres, which causes a dark
brown crisscross venation pattern that sometimes turn chlorotic; and the
spot form (SFNB), caused by P. teres f. maculata, which causes dark
brown circular or elliptical spots accompanied by chlorosis of the
surrounding leaf tissue (Khan and Tekauz 1982). Yield losses of 20 to 30
% in susceptible cultivars have been reported in western Canada (van den
Berg 1988) and up to 40 % in other parts of the world (Khan 1987). More
important than yield losses, the pathogen reduces thousand kernel
weight, plumpness and test weight, negatively affecting malting and feed
quality. The most effective and economical method to control this
disease is the use of resistant cultivars, however most commonly grown
barley cultivars are susceptible to most isolates of P. teres (Tekauz
1990, 2000). The variability observed in P. teres and failure to find
lines resistant to all isolates suggests breeding for resistance should
emphasize pyramiding resistance genes to develop broad-based durable
resistance. Molecular markers allow breeders to rapidly introgress
resistant genes into elite lines and to pyramid more than one resistant
gene into a cultivar. Molecular markers linked to net blotch resistance
in barley have been recently reported from Australia (Cakir et al. 2003,
Raman et al. 2003, Williams et al. 2003). There is need to determine
whether we can use Australian developed molecular markers in western
Canadian barley breeding programs. Thirty-nine barley lines were
screened with 6 NFNB (WRS102, WRS858, WRS1607, WRS1906, LO256, LO246)
and 4 SFNB (WRS857, WRS1881, LO233, LO231) isolates at the seedling
stage in the U of S, College of Agriculture Phytotron. Parents of
Australian barley mapping populations used to identify/validate net
blotch markers in Australia, western Canadian lines, US lines and one
Ethiopian accession and lines from International collections were
included. The majority of the Australian parent lines were susceptible
to western Canadian isolates. Australian \'R\' lines Franklin, Alexis,
Kaputar, Baudin, Hamelin were susceptible to the majority of the
isolates tested. However, Pompadour, Halcyon, Tilga and Chebec were
resistant to some isolates. Accession CI9214 showed the best overall
resistance to all isolates and several western Canadian lines/cultivars
viz. TR253, TR251, TR244, CDC Helgason and CDC McGwire were resistant to
the majority of the isolates. We will evaluate Pompadour/Stirling,
WPG8412/Stirling and Sloop/Halcyon populations to validate Australian
NFNB markers using western Canadian NFNB isolates. For SFNB marker
validation, potential populations are CI9214/Stirling, Keel/Gairdner,
Chebec/Harrington and Tilga/Tantangara.

##### Selected references:

Cakir, M. et al. 2003. *Australian J. Agric. Res.* 54:1369-1377.

Khan, T.N.1987. *Australian J. Agric. Res.* 38:671-689.

Raman, H. et al. 2003. *Australian J. Agric. Res.* 54:1359-1367.

Tekauz, A. 1990. *Canadian J. Plant Pathol.* 12:141-148.

Williams, K.J. et al. 2003. *Australian J. Agric. Res.* 54:1387-1394.

## Selection for improved scald resistance in the Crop Development Centre barley improvement program

### B.G. Rossnagel^1^, D. Voth^1^, T. Zatorski^1^, D.D. Orr^2^ and T.K. Turkington^2^

#### ^1^ Crop Development Centre, University of Saskatchewan, Saskatoon, SK, S7N 5A8 and ^2^ Agriculture and Agri-Food Canada, Lacombe Research Station, Lacombe, AB, T4L 1W1

Scald, caused by *Rhynchosporium secalis*, is a significant barley
disease in western Canada in most years, especially in the moist regions
of Alberta and north west Saskatchewan. Control depends primarily on
genetic resistance in varieties planted and cultural practices including
crop rotation and tillage to bury crop residue. Sources of resistance
have been identified; however, many resistance sources break down rather
quickly. Success in using the strategy of genetic resistance to control
scald has been hampered by evolution and pathogenic variability within
scald populations. Improved scald resistance is a part of the Crop
Development Centre (CDC) barley improvement program and involves
repeated testing of selected lines in a search for putative sources of
resistance as well as to monitor scald resistance inherent in the
program. Annual testing is conducted in collaboration with T.K.
Turkington et al, AAFC, Lacombe at screening nurseries at Lacombe and
Edmonton, Alberta. Scald reaction data has been collected at both
locations for more than 10 years. Data from 2000 through 2004 is
presented for a number of resistant CDC selections and six check
varieties. Intermediate resistance derived from CDC Dolly is holding but
appears to be less effective than it was initially. Resistance derived
from PC11 continues to be effective. Resistance tracing to Arizona
hulless waxy (AzHull) remains effective with a large number of
selections from crosses with that resistance currently being tested at
advanced levels in the CDC program. Lines derived from Senor demonstrate
moderately resistant reactions. Resistance from *Hordeum spontaneum* or
*H. bulbosum,* introduced to the CDC program via New Zealand accessions,
has endured well. Several CIMMYT lines, notably Calicuchima, 18 IBON-128
and 18 IBON-75 exhibit highly resistant reactions. Progeny of BT474, a
six row CDC breeding line with no apparent resistance source(s) in its
pedigree and now serving as a resistant nursery check, will be screened
for scald reaction in 2005 Alberta nurseries. Several selections from
crosses with BT474 have now reached advanced yield test stages of the
CDC program.

## Selection for improved FHB tolerance in the Crop Development Centre barley improvement program

### B. Rossnagel^1^, D. Voth^1^, T. Zatorski^1^, J. Tucker^2^, W. Legge^2^, and M. Savard^3^

#### ^1^ Crop Development Centre, University of Saskatchewan, Saskatoon, SK, S7N 5A8 ^2^ Agriculture and Agri-Food Canada, Brandon Research Station, Brandon, MB, R7A 5Y3 ^3^ Agriculture and Agri-Food Canada, Eastern Cereal and Oilseed Research Centre, Ottawa, ON K1A 0C6

Continued financial support by the WGRF, CWB and AAFC MII fund to
operate a collaborative Fusarium Head Blight (FHB) screening project,
initiated in 2000, at AAFC Brandon, attests to the serious impact of FHB
on both barley growers and users. As complete resistance has not been
observed in barley, genetic resistance from additive action of multiple
minor genes offers the greatest potential against FHB. The Crop
Development Centre (CDC) barley improvement program's participation in
this collaborative project was initially limited to searching for
putative sources of resistance for the breeding program and screening
existing breeding lines to select for improved reaction to FHB. Repeated
testing, 2001 to 2004, identified 9 hulled genotypes that consistently
demonstrate DON concentrations comparable to the resistant check CI4196.
Two of these lines, 2ND16092 (from NDSU) and HDE84194-622-1 (a Chinese
accession from Shanghai Academy) have been used as parents in the CDC
program on the basis of their FHB reaction. 114 selections from 7
populations derived from 2ND16092 and 65 selections from 2 populations
derived from HDE84194-622-1 will be tested in 2005 for FHB response as
Preliminary yield test entries. The remaining two-rowed hulled CDC lines
all share TR251 as a common parent. SB00106, the most promising in an
agronomic sense, also showed promise as an entry in the 2003 and 2004
North American Barley Scab Evaluation Nursery, where its DON level
across all sites was among the lowest. SB00106 was entered in the
Western Two Row Coop Test as TR04378 in 2004 and chosen to replace
CI4196 as the resistant check in the 2005 Brandon FHB nursery. The
larger proportion of hulless, 30 versus 9 hulled lines, repeatedly
selected and tested for low DON concentration, emphasize the
relationship between hull removal and DON accumulation. DON
concentrations for all selected hulless lines were lower than the
checks. The predominance of several parents (i.e. CI4196, CDC Freedom
and TR251) in the pedigrees of these selections indicate the heritable
nature of the low DON accumulation trait. SH00749, selected from an
early cross between CDC Freedom and CI4196 (population 99T511-03) was in
turn used as a parent to improve FHB tolerance. 115 selections from 4
populations derived from SH00749 will be tested in the 2005 FHB nursery
as pre-yield test entries. The FHB response of several of the best CDC
selections for lower DON accumulation, with few exceptions, seem to hold
up well in other FHB screening nurseries.

Dr. B.G. Rossnagel: rossnagel.brian@usask.ca

## An AFLP derived tightly linked marker for true loose smut resistance (*Un*8)

### Peter Eckstein, Donna Hay, Brian Rossnagel, and Graham Scoles

#### Department of Plant Sciences/Crop Development Centre, University of Saskatchewan, Saskatoon, SK, CANADA S7N 5A8

True loose smut resistance in barley is conferred by the single dominant
gene *Un*8, and is the major source of resistance in cultivars bred for
western Canada. Since smut is seed borne, conventional disease screening
involves the manual (hand held syringe) inoculation of 10 to 12 florets
per spike (line). Mature seed from inoculated plants is harvested and
grown to anthesis before plants can be evaluated for the presence of
smut, a process which needs to be repeated for putative resistant lines
because of potential escapes. Cost estimates for conventional screening
are near \$5 per line. This expense and the simple genetics of the
resistance make smut resistance an excellent candidate for
marker-assisted selection (MAS). PCR based markers for *Un*8 (Eckstein
*et* *al*., 2002) have been in routine use in the Crop Development
Centre barley breeding program for more than five years. While the
markers are robust and simple to use, the 6cM of genetic distance
(recombination) renders approximately 50% of crosses as monomorphic
(unscreenable). AFLP on bulked-segregant DNA samples has generated a
polymorphism that is more tightly linked with the resistance in 149 DH
lines from the cross Harrington x TR306. AFLP primers E32-M58 amplify a
short fragment of DNA from the resistant DNA bulk and resistant parent
only. This fragment is consistently amplified from constituent lines of
the resistant DNA bulk, is absent from the susceptible lines, and
co-segregates appropriately with the disease reaction in all lines that
showed recombination with our previous marker. Efforts to convert the
AFLP polymorphism to a simple PCR-based marker are in progress to
simplify linkage analysis on an additional 367 lines from the same
population. The polymorphic fragment has been isolated, cloned and
sequenced, and consists of 40 bases of genomic sequence from which
allele specific PCR-based markers cannot be designed. A 1052bp fragment
containing the original 40 bases has been identified through anchored
PCR. This locus has been sequenced from resistant and susceptible
genotypes in order to design allele specific primers. Analysis of
sequence to date indicates that the locus likely has numerous locations
in the barley genome and efforts to identify nucleotide variation
between resistant and susceptible genotypes at the *Un*8 linked locus
continue. This tightly linked marker will be useful in reducing the
error percentage in MAS, and will increase the amount of breeding
material that can be evaluated through MAS. Linkage estimates obtained
from the larger population will form the basis for closer examination of
the locus and perhaps the isolation of the gene itself. The eventual
isolation of the *Un*8 gene will allow for the characterization of
previously un-phenotyped materials such as can be done with the *Rpg*1
resistance gene (Brueggeman *et al*., 2002) and markers (Eckstein *et*
*al*., 2003).

##### References

# Brueggeman, R., Rostoks, N., Kudrna, D., Kilian, A., Han, J., Druka, A., Steffenson, B., and A. Kleinhofs. 2002. Proc. Natl. Acad. Sci. USA 99:9328-9333. {#brueggeman-r.-rostoks-n.-kudrna-d.-kilian-a.-han-j.-druka-a.-steffenson-b.-and-a.-kleinhofs.-2002.-proc.-natl.-acad.-sci.-usa-999328-9333. .References}

# Eckstein, P.E., Krasichynska, N., Voth, D., Duncan, S., Rossnagel, B.G., and G.J. Scoles. 2002. Can. J. Plant Pathol. 24:46-53. {#eckstein-p.e.-krasichynska-n.-voth-d.-duncan-s.-rossnagel-b.g.-and-g.j.-scoles.-2002.-can.-j.-plant-pathol.-2446-53. .References}

# Eckstein, P., Rossnagel, B., and G. Scoles. 2003. Barley Genetics Newsletter 33:7-11.  {#eckstein-p.-rossnagel-b.-and-g.-scoles.-2003.-barley-genetics-newsletter-337-11. .References}

## Cytological karyotyping of *Pyrenophora teres*

### A.D. Beattie, G.J. Scoles, B.G. Rossnagel

#### University of Saskatchewan/Crop Development Center, 51 Campus Drive, Saskatoon, SK S7N 5A8

Net blotch, caused by the fungal pathogen *Pyrenophora teres*, is a
common disease of barley that adversely affects seed quality
characteristics like plumpness and test weight. Despite its importance,
there is limited information available about the *P. teres* genome.
Karyotype analysis of this pathogen was initiated to address this and to
provide a foundation for further genetic work including the creation of
a molecular linkage map.

One technique available for karyotyping fungal pathogens is the germ
tube burst method (GTBM) which allows cytogenetic analysis by releasing
mitotic metaphase chromosomes from the actively growing tip of conidial
germ tubes. Previous cytological work with fungi focused on meiotic
stage chromosomes present during sexual reproduction, however, the
sexual stage of many important phytopathogenic fungi is either unknown
or difficult to induce in the lab, restricting the use of this method.
Pulsed field gel electrophoresis (PFGE) is another popular method of
karyotyping fungi which is useful for estimating the molecular weights
of chromosomes, but tends to underestimate chromosome numbers due to the
inability to resolve chromosomes of similar size. The combination of the
GTBM and PFGE make a powerful tool for karyotype analysis.

The GTBM was applied to four isolates of *P. teres*. A time course
analysis was conducted to determine the conidial germination rate and
the point at which the maximum number of germ tube nuclei were in
metaphase. Germination began after 30 min and was near 100% by 120 min.
The proportion of metaphase stage nuclei in germlings reached a maximum
of 15% at regular intervals of 60-70 min. Culture media was amended with
hydroxyurea to attempt to synchronize mitosis and increase the
proportion of metaphase nuclei, but only a marginal increase was
observed. Cells within the germling contained an average of seven nuclei
and the nuclei within each cell appeared to be at the same mitotic
stage. These observations were not significantly different between the
four isolates studied. Nine chromosomes were observed for each isolate
after making a minimum of 20 chromosome counts. Observations on
chromosome size were recorded. This karyotype analysis solidifies the
only previous estimation of chromosome number in *P. teres*, made using
PFGE. In that study, only six bands could be resolved, but densitometric
analysis of the larger, unresolved bands led to a chromosome number
estimate of nine. This study shows the power of the GTBM to accurately
determine karyotypes in fungi.

Corresponding author: adb164@duke.usask.ca (Aaron Beattie)

## A molecular linkage map of *Pyrenophora teres*

### A.D. Beattie, G.J. Scoles, B.G. Rossnagel

#### University of Saskatchewan/Crop Development Center, 51 Campus Drive, Saskatoon, SK S7N 5A8

Genetic linkage maps provide basic information about a species' genome
organisation and are important tools required for positional cloning of
genes. The rice blast pathogen, Magnaportha grisea, provides a good
example of how a linkage map of this fungus has allowed map-based
cloning of several avirulence genes. The goal of this project was to
construct a linkage map of the barley net blotch pathogen, Pyrenophora
teres, that will support future mapping studies.

A mapping population of 80 single ascospore progeny were isolated from a
cross between parental isolates WRS 1906 x WRS 1607. The parents were
screened with 144 AFLP primer combinations from which 23 primer pairs
were selected to screen across the entire population. Because WRS 1906
is avirulent on the barley variety \'Heartland\' while WRS 1607 is
highly virulent, the population was also evaluated for virulence on
Heartland and segregated 38 avirulent: 42 virulent (χ2 = 0.2, P = 0.70).
This suggested a single gene controlled the avirulent phenotype and this
locus was placed on the linkage map. Finally, the mating type
(MAT1/MAT2) locus was mapped on the population using a set of PCR
primers specific to this locus. The map consists of 110 unique loci
distributed over 18 linkage groups. Only eight (7.3%) markers displayed
a Mendelian segregation ratio different from 1:1. The total map length
is approximately 650 cM.

The present chromosome number estimate for P. teres is nine based on
pulsed field gel electrophoresis (PFGE) and cytological observations.
This clearly indicates that many of the linkage groups represent common
chromosomes. Currently markers from each linkage group are being
hybridized to PFGE-separated chromosomes in order to assign linkage
groups to specific chromosomes. Mapping of the chromosome telomeres has
also been initiated to determine how fully the current map represents
the genome and to better define the region around the avr locus near the
terminus of linkage group 6.

Corresponding author: adb164@duke.usask.ca (Aaron Beattie)

## The barley stem rust resistance gene *Rpg5* encodes NBS-LRR and protein kinase domains in a single gene

### R. Brueggeman^1^, T. Drader^1^, A. Druka^2^, T. Cavileer^3^, B. Steffenson^4^, J. Nirmala^1^, H. Bennypaul^1^, K. Gill^1^ and A. Kleinhofs^1,5^

#### ^1^Washington State University, Crop and Soil Science, Pullman, WA 99163, USA ^2^Scottish Crop Research Institute, Invergowrie, Dundee DD2 5DA, Scotland, UK ^3^University of Idaho, Dept. of Biological Sciences, Moscow, ID 83844, USA ^4^University of Minnesota, Department of Plant Pathology, St. Paul, MN 55108-6030, USA ^5^Washington State University, School of Molecular Biosciences, Pullman, WA 99163, USA

The biotrophic fungus *Puccinia graminis* causes stem rust of barley.
Several major genes for resistance (i.e. *Rpg1-Rpg5*) have been
described in barley. To understand the molecular basis of stem rust
resistance, we have focused on the isolation and characterization of the
genes involved in the incompatible interactions between barley and
*Puccinia graminis*.

The *Rpg1* gene was recently cloned and predicted to code for a
receptor-like kinase with dual kinase domains (Brueggeman et. al., PNAS
99:9328, '02), representing a novel class of plant disease resistance
genes. A genetic locus believed to contain the barley stem rust
resistance genes *Rpg5* and *rpg4* was delimited genetically to two BAC
clones and completely sequenced. The *Rpg5* locus confers resistance to
the rye stem rust pathogen *Puccinia graminis* f. sp. *secalis*, isolate
92-MN-90 and the *rpg4* locus confers resistance to *P. graminis* f.
sp*. tritici* pathotype, Pgt-QCC. Annotation of the BAC sequences
revealed several candidate resistance genes. Allele sequencing from
resistant and susceptible cultivars and recombinant lines resulted in a
single candidate *Rpg5* gene. The *Rpg5* gene was confirmed by allele
sequencing and it also appears to be required for *rpg4*-mediated
resistance. This was indicated by the presence of recombinants resistant
to isolate 92-MN-90 (*Rpg5*), but not to QCC (*rpg4*). Recombinants
resistant to QCC, but susceptible to 92-MN-90 were never isolated among
over 5,000 gametes examined. *In-silico* translation of the *Rpg5*
sequence from the resistant line Q21861 revealed a protein containing
NBS-LRR and protein kinase domains, all in one gene. Several cases are
known in the literature where an NBS-LRR gene and a protein kinase gene
are required for resistance to a pathogen, but this is the first case
where all three domains are encoded by a single gene. Further validation
of the gene is underway using a viral induced gene silencing approach as
well as complementation by *Agrobacterium* mediated transformation.
Characterization and validation of this gene will be presented and
possible mechanism of resistance discussed.

Robert Brueggeman: bigbass@wsu.edu

## *In vitro* selection for pre-screening barley for resistance to *Fusarium* head blight

### Kumar, K. (1), Xi, K. (1), Helm, J.H. (2), Turkington, T.K. (3), and Jennifer Zantinge (1) 

#### 1: Field Crop Development Centre, Alberta Agriculture, Food and Rural Development (AAFRD), 6000 C & E Trail, Lacombe, AB T4L 1W1 2: Field Crop Development Centre, AAFRD, 5030-50 Street, Lacombe, AB T4L 1W8 3: Lacombe Research Centre, Agriculture and Agri-Food Canada, 6000 C & E Trail, Lacombe, AB T4L 1W1

Cultivation of susceptible wheat and barley cultivars has resulted in
FHB epidemics in the Midwest USA and Manitoba. The use of barley
cultivars with genetic resistance to FHB is the most cost effective and
environmentally sound means to manage this disease. However, the limited
occurrence of FHB caused by *Fusarium graminearum* Schwable in Alberta,
especially in the central and northern regions, requires developing
methodology to screen resistance under controlled conditions. Barley
genotypes (*Hordeum vulgare* L.) with known field reactions to fusarium
head blight (FHB) were used for *in vitro* ground grain and leaf
detached assays. In the ground gain assay, the FHB reaction of the
genotypes was measured based on the extent of mycelial growth of *F.
graminearum* (PW027) on a mixture of agar and ground grain at room
temperature (23±1°C). In addition, the deoxynivalenol (DON) level in a
mixture of *F. graminearum* biomass and ground grain of each line was
determined using an ELISA-based assay. *Fusarium graminearum* showed
larger colony diameters (mm) for susceptible 'AC Lacombe' and 'Stander'
compared with resistant genotypes, 'Chevron' H94051001, I92130, and
H93120, except for resistant CI4196. *Fusarium graminearum* produced
more DON on susceptible 'AC Lacombe' compared to resistant 'Chevron',
I92130, and H93120, except for resistant CI4196 and H94051001. Larger
colony diameters and higher DON content were also observed for
susceptible genotypes compared with resistant ones in a repeated test
with another 16 barley genotypes. Detached leaves of six barley
genotypes were inoculated with single isolates of *F. graminearum*
(PW027) and *F. culmorum* (PW0T6) for evaluation of partial disease
resistant (PDR) components at 10°C and 23±1°C, and DON content at
23±1°C. Both isolates were pathogenic at 23±1°C with shorter incubation
period and greater lesion sizes on detached leaves compared to 10°C.
PW027 was more virulent than PW0T6 at both temperatures resulting in
shorter incubation and larger lesions for all barley genotypes but not
for spore production. *Fusarium graminearum* appeared to better
differentiate between resistant and susceptible barley genotypes at
23±1°C compared with *F. culmorum*. However, there was no clear pattern
in DON content between resistant/tolerant and susceptible genotype. The
results of *in vitro* evaluations tended to agree with previous field
reactions in terms of FHB reaction and DON level of the genotypes. Thus,
both *in vitro* assays may be alternate selection methodologies for FHB
resistance. The detached leaf assay has the advantage of measuring
specific disease components, allowing elucidation of the potential
nature and genetic components of resistance to FHB operating for this
assay. The potential of screening barley embryos of different genotypes
against DON as a method of identifying resistant genotypes was also
evaluated. In a preliminary test, 20 days old embryos of susceptible 'AC
Lacombe' and resistant H94051001 were evaluated in Murashige Skoog media
amended with different concentrations of DON. DON was found to
considerably reduce the embryo weight of 'AC Lacombe' compared with that
of H94051001. Further embryo experiments are being carried out, with
screening of more barley genotypes against different DON concentrations.

## Diversification strategies for barley disease management in Alberta

### Turkington, T.K. (1), Xi, K. (2), Clayton, G.W. (1), Harker, K.N. (1), O'Donovan, J.G. (1), and Lupwayi, N. (1)

#### 1: Lacombe Research Centre/Beaverlodge Research Farm, Agriculture and Agri-Food Canada, Lacombe, AB, T4L 1W1; 2: Field Crop Development Centre, Alberta Agriculture, Food and Rural Development, c/o Lacombe Research Centre, Agriculture and Agri-Food Canada, Lacombe, AB, T4L 1W1

Barley is an important feed grain crop in Alberta that accounts for
40-50% of Canadian production. To ensure a constant supply of feed many
farmers often grow barley continuously for several years; however,
continuous barley production leads to a build-up of plant diseases and a
general reduction in yield potential over the long-term. A three-year
experiment was established at Lacombe in 1998 to determine if barley
cultivar rotation could be used to reduce the impact of leaf diseases,
while maintaining crop productivity. Treatments consisted of various
sequences of two cultivars with varying degrees of scald and net blotch
resistance, 'Kasota', and 'AC Lacombe'; a previously scald-resistant
cultivar 'CDC Earl'; a susceptible check, 'Harrington'; and a non-host,
triticale cultivar 'Wapiti'. Rotations were established in 1998 with
comparisons being made in 1999 and 2000. In 1999, significant rotation
differences occurred for scald and net blotch severity, with disease
severity usually highest and yield lowest when a barley cultivar was
grown on its own residue, especially for cultivars other than 'Kasota'.
Statistical analysis using contrasts indicated that yield and kernel
weight were lower, while scald and net blotch levels were higher for
barley cultivar monoculture compared with barley cultivar rotation. In
2000, when a barley cultivar was grown on its own residue, scald
severity was usually highest compared to barley cultivar rotation. A
similar trend was also observed for net blotch, especially for
'Harrington' and 'AC Lacombe'. Poor stand establishment in some plots
precluded the detection of yield differences among some treatments in
2000. Contrasts for 2000 indicated that higher levels of scald and net
blotch, and decreased kernel and test weights occurred for barley
cultivar monoculture compared to barley cultivar rotation. In both 1999
and 2000, barley planted on triticale residue generally had the highest
yield, kernel weight and test weight, while having the lowest disease
levels compared to planting barley on barley residue. Barley cultivar
rotation can be a potential short-term strategy to help reduce leaf
disease levels and maintain crop productivity for Alberta barley
producers where crop rotation options are limited due to feed
requirements or market factors. However, continuous production of
barley, even utilizing cultivar rotation, will not provide long-term
sustainable leaf disease management, especially for scald. The summer of
2004 was the third year of a separate trial that was established to look
the interactive effects of agronomic factors such as rotational
diversity, seeding rate, and time of silage removal on crop health,
competitiveness, disease levels, productivity and quality in a cereal
silage production system. In 2004, all plots except for the continuous
triticale were seeded to the barley cultivar 'Seebe'. The rotation
treatments of 'CDC Helgason' barley/'Pronghorn' triticale/'Seebe' and
'Pronghorn'/'AC Mustang' oats/'Seebe' had significantly greater
emergence compared to the continuous 'Seebe' treatment, which had the
lowest emergence. The remaining rotation treatments had intermediate
emergence and were not significantly different from the continuous
'Seebe' rotation. Overall, silage yield on a dry weight basis was
highest for the 'Pronghorn'/'AC Mustang'/'Seebe' rotation (8060 kg/ha),
intermediate for the 'CDC Helgason'/'AC Mustang'/'Seebe' rotation (7822
kg/ha) and lowest for the remaining rotations (7005-7279 kg/ha, LSD =
400 kg/ha). The spot-form of net blotch was the main leaf disease
present and it was highest for the continuous 'Seebe' rotation (13.8%
leaf area diseased on the flag -2 leaf) and lowest for the
'Pronghorn'/'AC Mustang'/'Seebe' rotation (5.9%) with the other
rotations having intermediate disease levels (8.2-11.7%, LSD = 2.1).
Rotation had a significant influence on root mass assessed in the fall.
Root mass was highest for the 'Pronghorn'/'AC Mustang'/'Seebe' rotation
(42.4 g/2 m length of row), lowest for the continuous 'Seebe' rotation
(29.1 g), and intermediate for the remaining rotation treatments
(34.4-37.0 g, LSD = 8.2 g). Rotation treatments had an impact on silage
yield and this appeared to be related to crop health as indicated by
leaf disease levels and root mass measurements. A second 3-year cycle of
this trial is being repeated starting in 2005.

E-mail: turkingtonk@agr.gc.ca

## Resistance of western Canadian barley genotypes to scald in Alberta

### K. Xi (1), T.K. Turkington (2) and C. Bos (1)

#### 1: Field Crop Development Centre, Alberta Agriculture, Food and Rural Development, c/o Lacombe Research Centre, 6000 C&E Trail, Lacombe, AB Canada T4L 1W1 2: Agriculture and Agri-Food Canada, Lacombe Research Centre, 6000 C & E Trail, Lacombe, AB Canada T4L 1W1

Barley (*Hordeum vulgare* L.) production in Alberta has averaged around
6 million metric tonnes from 2 million hectares annually from 1995 to
1999, and accounted for close to 50% of the total barley production in
western Canada. Scald caused by *Rhynchosporium secalis* J. J. Davis is
one of the major foliar diseases causing significant yield loss in
Alberta as a result of intensive barley production, and cool and wet
environmental conditions that favor disease development. In the current
study, nine barley differentials were grown up in hill plots at multiple
sites to monitor scald development during 1997 -- 1999 (Period 1),
twelve during 2000 -- 2001 (Period 2), and twelve during 2003 -- 2004
(Period 3). Thirty-eight genotypes and commercial cultivars with
different levels of resistance were also evaluated for scald reaction
from 2003 to 2004 at these sites. Differentials Abyssinian, Atlas, Atlas
46, Atlas 68, Hudson, Osiris, Kitchin and Turk were resistant to scald
at all sites, other differentials including Brier, La Mestita, Modoc and
Trebi showed intermediate levels of scald, indicating that the majority
of major genes for scald resistance are effective against *R. secalis*
pathotypes in Alberta. The differentials showed highly significant
correlations in scald severity among all three periods. Period 1 and
Period 2 (*R*^2^=0.86\*\*) were, however, more closely correlated,
compared with Period 1 and Period 3 (*R*^2^=0.63\*\*), suggesting the
development of considerable variability in *R. secalis* in response to
major genes for resistance during 1997 to 2004 in Alberta. Analysis from
ten station-years of data classified thirty-eight genotypes and
cultivars into three major clusters corresponding to scald reaction:
resistant, intermediate and susceptible. Those in Cluster I that were
resistant included AC Stacey, Kasota, Mahigan, Manny, Niobe, Seebe and
five other genotypes from the Field Crop Development Centre. Eighteen
commercial cultivars in Cluster II were intermediate in scald reaction
and these included AC Lacombe, CDC Dolly, CDC Earl, CDC Guardian, CDC
Kendall, Falcon, Nisku and Peregrine. The three susceptible barley
cultivars including Harrington were classified into Cluster III with
several malting cultivars including AC Metcalfe, CDC Sisler and Excel,
and three feed barleys, AC Bacon, AC Rosser and CDC Candle. The number
of cultivars that were classified to be resistant was up from previous
studies. A substantial number of feed cultivars were classified to be
intermediate in resistance in the present study due partly to the
overall moderate level of scald severity during 2003 -- 2004 in Alberta.
Changes in *R. secalis* pathotypes and conducive weather conditions may
increase severity, resulting in breaking down of resistance, as
demonstrated in previous study. Given the fact that no malting cultivar
was resistant and only one was classified as intermediate in scald
reaction, there is a need to incorporate scald resistance genes into
malting cultivars for production in Alberta.

## Mapping and molecular marker development of scald resistance in 'Seebe' barley

### Zantinge, J.L.\*, J.H. Helm, Z. Hartman, J.B. Russell, and K. Xi

#### Field Crop Development Centre, Alberta Agriculture, Food and Rural Development, 5030 -- 50th Street, Lacombe, AB T4L 1W8, Canada. Web Site: <http://www1.agric.gov.ab.ca/app21/rtw/selsubj.jsp> 

Scald (*Rhynchosporium secalis*) of barley is prevalent in central
Alberta, Canada and causes considerable yield and quality losses. Scald
can rapidly change in pathotype composition and frequency, thereby
making it difficult to develop durable scald resistance in barley.
Previous studies have shown that the cultivar 'Seebe' carries durable
genetic resistance, however, barley breeders have found this trait
difficult to transfer into new barley lines. Therefore, we are trying to
develop molecular markers for scald resistance from 'Seebe'. Recombinant
inbred lines were developed from the cross of 'Harrington' (scald
susceptible) and 'Seebe' (scald resistant). Progeny of about 175
individual F~2~ seedlings were advanced by single-seed descent to the
F~8~ generation. Disease resistance to a major scald race was phenotyped
at the seedling stage in a greenhouse. By utilizing bulked segregant
analysis (BSA), resistant and susceptible pooled populations were
compared by AFLP analysis. A total of 255 AFLP primer combinations were
used to analyze the genetic population and several *Eco*RI-*Mse*I and
*Pst*I-*Mse*I fragments were found linked to scald disease resistance.
We are also analyzing this population with SSR markers. Our goal is to
map and identify molecular markers flanking the genes contributing to
scald resistance in 'Seebe'.

**Key words:** scald resistance, marker development, *Hordeum vulgare*,
barley

\*Corresponding author: jennifer.zantinge@gov.ab.ca

## Breeding for multiple disease and multiple gene resistance in barley

### James H. Helm^1^, H. Vivar^2^, F. Capettini^2^, K. Xi^1^, P. Juskiw^1^, and J. Zantinge^1^ 

#### ^1^ Alberta Agriculture, Food and Rural Development; Field Crop Development Centre, Lacombe Alberta ^2^ ICARDA/ CIMMYT, Texcoco, Mexico 

Combining genes for disease resistance is very difficult, as most
breeding programs can only test for the diseases present at their
breeding sites. In addition, pyramiding genes or developing multiple
gene resistance is difficult to detect when testing at only one
location. Multiple disease and gene resistance involves breeding against
more than one pathogen and more than one gene per pathogen. Each
pathogen may have several races that are able to attack varieties and
render a resistant variety ineffective in a short period of time,
presenting a significant challenge to plant breeders.

Over the years the ICARDA/CIMMYT barley program in Mexico has given us
an excellent opportunity to screen for multiple gene resistance for
Scald in barley and at the same time look at multiple disease resistance
to Stripe Rust, Barley Yellow Dwarf Virus, Leaf Rust, and Fusarium Head
Blight (FHB). In Canada not only have we screened for Scald, but have
also screened for Loose Smut and Covered Smuts as well as Net Blotch
(net and spot forms), Spot Blotch, and FHB. Over the last 5 years we
have screened over 2000 breeding lines at 4 locations in Canada and 3
locations in Mexico. New combinations of resistance genes have been
found with some lines containing genes for resistance to 5 and 6
diseases. We found multiple gene combinations for scald resistance that
have 3 or more genes and should give durable resistance to this disease
in both countries. In order to classify breeding lines according to
resistance gene combinations, we are currently analyzing overall
similarity computed from multivariate disease resistance data and
matching it to the pedigree.

The best lines will be used in the breeding program in order to rapidly
incorporate even greater disease resistance into new varieties for
Alberta producers. We will also develop several populations to begin the
process of mapping on as many of these genes as possible. Continuation
of this research is necessary to anticipate and cope with the changes in
disease problems likely to occur in the future. Up to this point in
time, stripe rust has not been a problem in Alberta on barley; however,
in 2004 this disease was found on barley at Olds, Trochu, Calmar and
Lacombe. If this disease continues its move north it will be devastating
to Alberta's barley crop. FHB also is not presently a problem in Alberta
but seems to be moving west. FHB has cost the barley industry millions
of dollars in the Midwest in the United States and in Manitoba in
Canada.

### 

## Variation in virulence among net blotch isolates infecting barley

### Linnea G Skoglund

#### Busch Agricultural Resources, Inc. 3515 Richards Lake Road, Fort Collins, CO 80525

*Pyrenophera teres*, causal agent of net blotch of barley, is widely
distributed throughout North American production areas. Previously,
researchers have shown variation in virulence of isolates within and
between locations. The purpose of this research was to continue
monitoring diversity of isolates in locations of interest to the BARI
barley breeding program. Isolates were collected randomly from
symptomatic plants from 2000 to 2004. A set of 25 differential varieties
was developed based on work of other researchers. Differentials were
planted in Rootrainers containing Metromix 200 and grown in the
greenhouse. Isolates were grown on V8 Juice agar at 18C with 12 hr light
per day for 7 days. Plants were inoculated at the 2-leaf stage with a
suspension of a single isolate, incubated in the dark at 18C and 100% RH
for 24 hr, and returned to the greenhouse. Plants were evaluated for
disease on a 0 (R) to 9 (S) scale 7-10 days later. To date, 16 isolates
have been characterized on 25 differential varieties. Isolates ranged
from the least virulent (PT01/6 from Bottineau, ND), infecting 5
differentials, to the most virulent (PT04/14 from Minot, ND), infecting
20. Of these 25 differentials, only 2 (9839 and CIHO 5822) were
resistant to all isolates and 3 (Alexis, Bonanza and Klages) were
susceptible to all isolates. There was no correlation of virulence to
year or location. This study will continue in order to assist breeding
programs developing resistance to net blotch.

Corresponding author: linnea.skoglund@anheuser-busch.com

## Assessment of artificial inoculation methods and deoxynivalenol levels in barley lines representing various candidate sources of resistance to *Fusarium* head blight

### Geddes, J. (1), Eudes, F. (1), Legge, B. (2), Tucker, J. (2)

#### 1: Agriculture and Agri-Food Canada, Lethbridge, AB T1J 4B1

#### 2: Agriculture and Agri-Food Canada, Brandon, MB R7A 5Y3

*Fusarium* Head Blight (FHB) is one of the most devastating diseases
affecting the production of barley and other cereal grains throughout
Canada and the world. FHB infection results in drastic decreases in crop
yield and severe reduction in grain quality. The most common species
causing FHB in North America is *Fusarium graminearum* (Schwab). The
fungus produces mycelical extensions that rapidly spread among the
florets, leaving shrivelled kernels and floral pieces covered with a
pink or white film of mycelium displaying elevated deoxynivalenol (DON)
levels. 19 barley lines representing FHB candidate resistance and
susceptible sources were point inoculated or spray inoculated in the
greenhouse with 40,000M/ml of *F. graminearum* macroconidia. Following
inoculation barley plants were kept at 24°C and 95% humidity for 3 days
to favour disease establishment and were then returned to normal growing
conditions at 21°C without humidity control (45%). 18 days
post-inoculation heads were collected and spikelets displaying symptoms
of FHB infection were rated for progression of the disease. These lines
were also evaluated in the Brandon, MB nursery from 2000-2004. The
number of discoloured spikelets produced by point inoculation, indoor
spray inoculation, and disease and DON assessment were compared for each
of the 19 lines. Barley lines representing candidate sources of FHB
resistance or susceptible FHB sources showed varying degrees of symptoms
due to fungal infection in the three tests. Barley lines representing
more or intermediate FHB resistant sources showed consistent levels of
resistance in the three tests. Artificial inoculation methods and DON
quantification enable us to rate the level of resistance of the barley
lines, with confidence. A higher level of FHB resistance will guarantee
lower risks for the farmer associated with crop losses due to reduced
grain yield, low quality grain, and DON contamination.

## Reactions of barley lines to leaf rust, caused by *Puccinia hordei*

### Y. Sun (1), J. D. Franckowiak (2), and, S. M. Neate (1)

#### (1) Department of Plant Pathology, North Dakota State University, Fargo, ND 58105; (2) Department of plant sciences, North Dakota State University, Fargo, ND 58105 USA. 

Resistant barley cultivars are the most economical and efficient means
of controlling leaf rust caused by *Puccinia hordei* G. Otth. However,
changing virulence in *P. hordei* has rendered ineffective many of the
known resistance genes. In this study, isolates 'Race 8', '90-3',
'90-15', '89-3', and 'Neth 202' of *P. hordei* were used in the
greenhouse to differentiate resistance genes in a collection of 82
selected barley lines. These lines exhibited resistance in the previous
tests against isolate 'Race 8' at the seedling stage. Barley line
'C2-02-134-2-2' exhibited low reaction types to all the tested leaf rust
isolates, suggesting that in addition to an already introduced
resistance gene, *Rph15*, it possesses one or more new resistance genes.
The F~2~ population of a cross made between 'C2-02-134-2-2' and a
susceptible line 'ZA47' which was challenged with isolate 'Race 8',
segregated into a 15:1 resistant to susceptible ratio (*X^2^*=0.853)
based on disease reaction. The 15:1 segregation ratio indicates that
'C2-02-134-2-2' possesses two genes, *Rph15* and a new single dominant
resistance gene. In the F~2~, the *Rph15* phenotype (00;) was separated
from the new resistance gene phenotype (0;12-). To isolate the new gene
from *Rph15*, the 10 F~2~ plants bearing the new phenotype were
transplanted and selfed and the F~3~ will be screened for homogeneity of
disease reaction. The identification of the new resistance gene(s) and
incorporation of them into barley cultivars will add in reducing yield
losses due to leaf rust.

## The differences in fermentable carbohydrates of major Canadian malting barley varieties and their effects on fermentation 

### Yueshu Li^1^, Rob McCraig^1^, Ken Sawatzky^1^, Aleks Egi^1^ and Michael Edney^2^

#### 1. Canadian Malting Barley Technical Centre, Winnipeg, R3C 3G7 Canada 2. Grain Research Laboratory of the Canadian Grain Commission

##### Abstract

The fermentable carbohydrates of the major Canadian malting barley
varieties were monitored during malting and brewing processes. It was
observed that the fermentable carbohydrate compositions of the congress
wort were varietal dependent, while the fermentable carbohydrate
compositions of the finished wort were both varietal and mashing
condition dependent. The overall malt modification affected malt
carbohydrate composition, and malt's brewing performance and final beer
quality.

## NanoMash: A novel procedure for research mashing of limited-quantity barley malts

### Laurie A. Marinac and Mark R. Schmitt\*

#### USDA Agricultural Research Service, Cereal Crops Research Unit, 501 Walnut St., Madison, WI 53726

The ASBC Malt-4 extract analysis method (Congress Mash) provides a
standard set of conditions for generating an unhopped wort commonly used
to evaluate the malting quality performance of experimental malts.
However, the method requires specialized instrumentation and relatively
large quantities of malt. These requirements prevent researchers without
access to the specialized instrumentation and particularly those with
limited sample availability from generating Congress worts for malting
quality analysis and other research uses. Use of a commercially
available device allows agitation of small volume samples through
orbital mixing while heating or cooling them using a Peltier temperature
controlled block. Controlling the device to follow the Congress mash
temperature profile offers the possibility of conducting a mashing cycle
at a significantly reduced scale. Adaptation of standard ASBC wort
analysis methodology to microtiter plate and other reduced-scale
methodology allows provision of several key parameters for worts
generated from size-limited samples. While not intended to supplant
standard ASBC methodology, this small-scale mashing protocol
significantly lowers the sample requirements and extends the potential
for malt analysis to research programs where it may not have been
previously feasible.

\*Corresponding author: 608-262-4480, markschmitt@wisc.edu

## Comparison of hull peeling resistance of barley and malt in western Canadian two-row barley lines

### W.G. Legge^1^, J.S. Noll^2^, and B.G. Rossnagel^3^

#### ^1^Agriculture and Agri-Food Canada, Research Centre, P.O. Box 1000A, R.R. #3, Brandon, Manitoba R7A 5Y3; ^2^Agriculture and Agri-Food Canada, Cereal Research Centre, 195 Dafoe Road, Winnipeg, Manitoba R3T 2M9; ^3^Crop Development Centre, University of Saskatchewan, 51 Campus Drive, Saskatoon, Saskatchewan S7N 5A8.

Hull peeling resistance in barley and malt is desired by the malting and
brewing industry. A group of 15 two-row malting barley (*Hordeum
vulgare* L.) lines and one feed cultivar were used to compare these
traits in Canadian two-row barley and determine their relationship to
other agronomic traits, including grain yield, kernel plumpness, test
weight and kernel weight. Barley samples from standard yield test plots
at two sites in each of Manitoba and Saskatchewan during 1999 and 2000
were evaluated for % hull peeling on a weight basis direct from the plot
combine (RPWB) and after inducing peeling with an air-blast de-huller
(APWB). Micromalted samples were evaluated for % hull peeling of the
malt "as is" (RPWM) and after air-blast de-hulling (APWM). Analyses of
variance over years and locations indicated that hull peeling of barley
and malt were strongly influenced by environmental factors, particularly
year. However, the single largest source of variance for RPWM and APWM
was the genotypic component, indicating that heritability and response
to selection would be higher in malt than barley. Genotypic effects were
lowest for RPWB, but still highly significant (P\<0.001).
Genotype-by-environment interactions were also highly significant, but
lower in magnitude. Manley, Stein and CDC Dolly were consistently lower
in all hull peeling traits, while TR251, CDC Unity, AC Oxbow and AC
Bountiful were significantly higher with TR244 always being the highest.
The relative ranking of the barley lines varied somewhat with CDC Unity,
AC Bountiful and Merit ranking higher for susceptibility to hull peeling
as malt than barley, while AC Metcalfe, Harrington and TR253 were ranked
less susceptible as malt. Among hull peeling traits, the highest
correlation of 0.95 (P\<0.001) was between RPWM and APWM, indicating
that RPWM would be adequate for determining hull peeling in malt. The
correlation between RPWB and APWB was also high at 0.92 (P\<0.001), but
RPWB was more variable due to its relatively low values averaging about
1.0% overall compared with 36.8% for APWB. Other correlations among hull
peeling traits ranged from 0.82 to 0.92. There were moderate negative
correlations between test weight and the hull peeling traits, ranging
from -0.55 (P\<0.05) to -0.69 (P\<0.01). Kernel plumpness, kernel weight
and yield were not significantly correlated with any of the hull peeling
traits. In conclusion, it should be possible to develop two-row malting
barley cultivars with acceptable kernel characteristics and improved
resistance to hull peeling of both barley and malt.

Corresponding author: blegge@agr.gc.ca

## Elimination of barley colour defects in Australia

### Glen Fox^1^, Maria Sulman^1^ and Kevin Young^2^

#### ^1^Department of Primary Industries & Fisheries, Toowoomba, Qld 4350 Australia ^2^ GxE Crop Research, PO Box 1704, Esperance, WA 6450, Australia

Grain colour defects including staining and black point have been a
problem for Australian barley growers for a number of years, resulting
in thousands of tonnes of malting barley being downgraded each year.
Over the last decade, research has been conducted investigating many
aspects of these disorders, including objective assessment, biochemical
evaluation, crop management, storage effects and resistance breeding. A
number of breeding lines have been identified with resistances to both
forms of colour defects. The black point tissue has been extracted with
high levels of phenolic acids (namely ferulic and coumaric acids) being
present suggesting the dark pigmentation may be a large polyphenolic
compound. Black pointed grain also has an impact on germination rate.
Markers for black point resistance coincide with markers for dormancy
and pre-harvest sprouting amylase. The effects on fungal contamination
results in a dark staining on the grain. A number of factors impact of
the degree of staining including pre-harvest rainfall, timing of the
rainfall event and pre-existing level of resistance. Husk content also
appears to have an impact on the final appearance of the grain. Markers
for husk content also coincide with genetic regions for dormancy and
pre-harvest sprouting. The improvement of grain quality at intake can be
delivered through two options; 1. the development of barley varieties
with resistance to these grain defects as resistance to black point and
staining has been shown to be heritable and 2. optimising early harvest
strategies and cool-air drying on-farm.

[glen.fox@dpi.qld.gov.au](../glen.fox@dpi.qld.gov.au)

## 

## Characterization of barley tissue-ubiquitous beta-amylase2

### Suzanne E. Clark^1^, Patrick M. Hayes^2^, and Cynthia A. Henson^1,3^

#### ^1^Dept. of Agronomy, University of Wisconsin-Madison, ^2^Dept. of Crop and Soil Science, Oregon State University, and ^3^UDA-ARS Cereal Crops Research Unit, Madison, WI

There are two barley β-amylases genes, encoding important starch
degrading enzymes. The endosperm-specific β-amylase (Bmy1), the more
abundant isozyme in cereal seeds, has been thoroughly characterized. The
lesser abundant β-amylase2 (Bmy2), has not been biochemically
characterized from any cereal seeds. Characterization of Bmy2 from two
commonly grown barley (*Hordeum vulgare* L.) cultivars, 'Morex' and
'Steptoe', was a major objective of this study. The bmy2 cDNAs were
sequence, expressed in *Escherichia coli*, and the recombinant enzymes
(rBmy2) characterized. The relative hydrolysis rates of various
a-D-glucans and the pH activity optima of 'Morex' and 'Steptoe' rBmy2s
were the same and not significantly different from barley rBmy1. The
'Morex' rBmy2 was 7^o^C more thermostable than the 'Steptoe' rBmy2,
determined by differences in their T50 values, and is more thermostable
than any reported wild type β-amylase1. Three amino acid differences
were identified between the two Bmy2 sequences and the contributions to
enzyme thermostability evaluated by site-directed mutagenesis.
Examination of mutant enzymes with one amino acid substitution revealed
that each of the three residues contributed \~3^o^C to the
thermostability of the 'Morex' wild type rBmy2. Mutant enzymes with two
amino acid substitutions contributed \~5.6^o^C and the triple amino acid
mutant enzyme contributed \~8.7^o^C to thermostability. To date, no
quantitative trait loci (QTL) for malting quality traits have been
associated with the bmy2 locus. Should an association be discovered, the
'Morex' bmy2 allele, containing D238, M337 and Q362, provides a discrete
signature of a thermostable β-amylase2 that could be targeted for marker
assisted selection.

Corresponding author: cahenson@wisc.edu

## Barley seed osmolyte concentration as an indicator of preharvest sprouting

### Cynthia A. Henson^1,2^, Stanley H. Duke^2^, Paul Schwarz^3^, Rich Horsley^3^, and Charles Karpelenia^1^

#### ^1^USDA-ARS Cereal Crops Research Unit, Madison, WI 53706, ^2^Department of Agronomy, University of Wisconsin, Madison, WI 53706, ^3^Department of Plant Sciences, North Dakota State University, Fargo ND 58105

This study was conducted to test the hypothesis that barley seed
osmolyte concentrations can be used as an indicator of preharvest
sprouting (PHS). Osmolyte concentrations from the 2002 Minnesota and
North Dakota crops were compared to pearling and other techniques for
assessment of PHS. Approximately 30% of the seed evaluated were
sprouted. Samples were evaluated for osmolyte concentrations, pearling,
and Stirring Number, while smaller subsets were evaluated using other
methodologies. Osmolyte concentrations correlated well with pearling
(r=0.822, P\<0.0001) and fluorescein dibutyrate staining (r=0.835,
P\<0.0001). a-Amylase activity correlated less positively with osmolyte
concentration (r=0.650, P\<0.0001). Stirring Number, Falling Number,
germination, and accelerated aging had weak negative correlations with
osmolyte concentration. It is concluded that seed osmolyte
concentration, a relatively simple assay, is a good indicator of PHS in
barley as assessed by the pearling method or fluorescein dibutyrate
staining.

Corresponding author: cahenson@wisc.edu

## 

## 

## Osmolyte concentration as an indicator of malt quality

### Cynthia A. Henson^1,2^, Stanley H. Duke^2^, and Charles Karpelenia^1^

#### ^1^USDA-ARS Cereal Crops Research Unit, Madison, WI 53706, ^2^Department of Agronomy, University of Wisconsin, Madison, WI 53706

This study was conducted to test the hypothesis that malt osmolyte
concentrations can be used as an indicator of barley malt quality. Seeds
of four 6-row and four 2-row genotypes were steeped for six days at 20C
for 6 days. At intervals of 24 h over the steeping regime green malt
from each cultivar was removed and tested for osmolyte concentration,
malt extract (ME), diastatic power (DP), α-amylase activity,
soluble/total protein (S/T), and β-glucan concentration. Osmolyte
concentrations increased most rapidly in days 1 through 3 of
germination. After 4 days osmolyte concentrations began to plateau.
Significant positive correlations were found for malt extract and
osmolyte concentrations in days 1 through 4 and day 6 (r=0.740 to 0.942,
P\<0.0001). Days 2 and 3 osmolyte concentrations correlated well with ME
for all days (r=0.740 to 0.942, P\<0.0001) and α-amylase activity for
day 2 (r=0.771, P\<0.0001). Day 2 osmolyte concentration correlated well
with days 2 through 6 for β-glucan concentration (r= -0.702 to -0.830,
P\<0.0001). No significant correlations were found for DP and osmolyte
concentrations on any day. These data indicate that osmolyte
concentrations at early time points in steeping are good indicators of
several measures of malt quality at later time points in steeping.

Corresponding author: cahenson@wisc.edu

## Relationships among malt fermentability and malt quality parameters under the influence of barley β-amylase heat stable allele

### Blanca Gómez\*^1^, Héctor Acevedo^2^, and Ana Clara López^3^

#### ^1^Malting Unit, Cereal Department, Laboratorio Tecnológico del Uruguay, Av. Italia 6201 CP 11500, Montevideo, Uruguay ^2^Quality Laboratory, Malteria Oriental S.A. Abrevadero 5525, Montevideo, Uruguay ^3^Biotechnology Department, Laboratorio Tecnológico del Uruguay, Av. Italia 6201 CP 11500, Montevideo, Uruguay.

Starch hydrolysis during germination is achieved by the action of four
major enzymes, -and β-amylase, limit dextrinase and -glucosidase. This
hydrolysis produces fermentable sugars required for yeast nutrition in
brewing. Fermentability is a critical quality parameter for brewing that
affects the level of alcohol produced by yeast, and is typically
assessed in malt extracts by determining the change in specific gravity
after small scale fermentation, referred to as apparent attenuation
limit. Diastatic power is a measure of the capacity of the malt to
degrade starch into fermentable sugars and is primarily determined by
β-amylase activity. Although diastatic power is a reasonable predictor
of fermentability, it does not always accurately estimate the level of
fermentable sugar generated during mashing or the subsequent
fermentability of the resultant wort. β-amylase is one of the major
proteins found in the starchy endosperm, which rapidly loses activity at
mashing temperatures above 55ºC. Two alleles with different β-amylase
enzyme thermostability are distinguished with the insertion/deletion of
a palindromic 126-pb sequence in intron III of *β-amy1* gene. Increased
thermostability results in more efficient starch degradation. Barley
cultivars with high thermostability β-amylase allele will achieve high
levels of fermentability without a good malt modification. On the other
hand, barley cultivars with low thermostability β-amylase allele will
achieve high levels of fermentability if they achieve high levels of
malt modification. The objective of this study was to determine the
influence of heat stable β-amylase allele in the relationship between
fermentability and quality parameters. We characterized 40 commercial
malting barley varieties from different countries. The malt quality data
used in the correlations studies come from the average of the results
obtained by 50 laboratories. The quality malt parameters analyzed were:
Fine Grind Extract, Total Nitrogen, Hartong 45, Diastatic Power, Wort
Viscosity, Alpha-Amylase, Friability, Free Amino Nitrogen, Final
Attenuation Apparent and Soluble β-Glucans. Significant correlations
were obtained from the statistical point of view, but values of R square
explained less than 20 % of the variability among the parameters. In
general, barley varieties that present high thermostability β-amylase
allele showed better qualitative levels of fermentability and malt
modification than the varieties with low thermostability β-amylase
allele. From these results we conclude that the fermentability is not
easily predicted from quality parameters, therefore the use of *β-amy1*
allele in barley breeding program is necessary to improve fermentability
levels in malting barley grains. More studies are needed to understand
all the properties that influence malt fermentability and how they
interact.

\*Corresponding author: bgomez@latu.org.uy

## Russian wheat aphid resistant barley -- cultivar and germplasm release

### D.W. Mornhinweg^1^, P.P. Bregitzer^2^, D.A. Obert^2^, F.B. Peairs^3^, D. Baltensperger^4^ and R. Hammon^3^

#### ^1^ USDA-ARS, Stillwater, OK, ^2^ USDA-ARS, Aberdeen, ID, ^3^ Colorado State University, ^4^ University of Nebraska

RWA continues to be a devastating pest of barley in the high and dry
areas of the Western U.S.A. Screening of the entire National Small
Grains Collection in Aberdeen Idaho by the USDA-ARS in Stillwater
identified 115 accessions with some level of resistance ranging from 2
to 6 on Webster's scale of 1 to 9 where 1 is immune and 9 is dead.
Resistant germplasm lines were developed from each accession and two of
these lines, STARS 9301B and STARS 9577B were released in 1993 and 1995
respectively. A long term prebreeding project was initiated at the
USDA-ARS in Stillwater to develop adapted germplasm lines by bringing
multiple sources of resistance into barley cultivars and elite lines of
both state and federal barley breeders across the country. These
breeders as well as extension personal from several states have been
involved in field testing of the 62 prebred germplasm lines now ready
for release. A detailed description of these lines and a time table for
their release will be presented. Along the way several feed barley
cultivars have also been developed. The first RWA-resistant barley
cultivar, Burton, has released by USDA-ARS in Aberdeen in conjunction
with USDA-ARS in Stillwater and several other cooperators. Burton, a
2-rowed, hulled, spring barley, has shown excellent performance in
irrigated and dryland areas both in the presence and absence of RWA.
Three, 2-rowed, spring, feed barleys, developed by the USDA-ARS in
Stillwater and Aberdeen and which are adapted to the extremely arid
conditions of the western high plains are currently in seed increase and
planned for release this fall. A new biotype, RWA2, identified in
Colorado in the summer of 2003, has been found to damage all currently
grown wheat cultivars developed for resistance to the original biotype,
RWA1. All germplasm lines and cultivars slated for release from this
program have been found to be resistant to RWA2 as well as RWA1.

Do. Mornhinweg@ars.usda.gov

## BARMS: A new relational database for barley breeding programs

### D. B. Cooper and Bruce Westlund

#### Busch Agricultural Resources Inc., Ft. Collins, CO U.S.A. 80524

Breeding programs generate large amounts of phenotypic data on parents,
segregating populations and derived experimental lines. There currently
are very few commercially available databases that are specifically
designed for use in plant breeding programs.

After conducting an initial study of existing database options that were
permitted on our corporate computer systems, it was decided that we
should develop a proprietary data solution based on the Oracle ^TM^
relational database software engine. The development process was done in
stages that were user tested and then refined to meet the requirement
standards and user acceptance. The Barley Ag. Research Management System
(BARMS) went live in February of 2004 and has routinely used by our
barley breeding programs since.

One of most difficult problems in interpretation of multi-location,
multi-year data from agricultural experiments is separating the relative
contributions of genetics (G), environment (E) and G x E interaction.
This can be especially critical when comparing 'un-balanced' data sets
from lines at different stages of trialing that have not been grown in
the same number of station x years. One partial solution to such data
sets is to adjust all observations relative to one or more check
cultivars. Phenotypic data is stored in BARMS in standard units of
measure (i.e. Yield in Bu/A, Plant height in cm) and can be queried in
that standard format or in alternate units of measure (i.e. Yield in Kg
/ ha, plant height in inches). Data for all traits can also be retrieved
on a 1-99 **RP** (Relative Phenotypic) scale where data is set relative
to three known check cultivars. The six-row breeding program uses Morex,
Robust and Legacy; the two-row breeding program uses Harrington, B1202
and Merit as the three comparator checks. Data points where an
experimental line appears in the same trial with all three of the
respective checks will be used to calculate a line RP for that trait on
a 1-99 scale. In this form environmental and G x E interactions are
minimized and the RP value represents a best linear unbiased estimate of
the true genotypic contribution. These RP scores can be used to evaluate
lines from un-balanced data sets on a more equitable basis.

We designed a **SELECTION MODULE** in BARMS that permits selections to
be made using both independent culling with upper and lower limits on
single trait values as well as index selection based on weighted sum of
squares of deviation of multiple trait values from defined targets. It
is believed that BARMS is the only software program to permit
simultaneous use of both selection protocols.

We developed a **CROSS COMPARISON MODULE** that permits multi-trait
evaluation of parental phenotypic data in all possible pair-wise
combinations as a predictor of future overall success of the resulting
progeny of a cross.

We automated our malt quality laboratory equipment to directly upload
evaluation results into the BARMS database. This has reduced the manual
entry time and increased accuracy of the micro-malting data coming from
our laboratory.

## Mapping and molecular marker development of seed dormancy in a barley population derived from 'Samson' barley

### J.L. Zantinge\*, J.M. Nyachiro, S. Chisholm, J.H. Helm, P.E. Juskiw and D.F. Salmon

#### Field Crop Development Centre, Alberta Agriculture, Food and Rural Development, 5030-50 Street, Lacombe, AB T4L 1W8, Telephone: (403) 782-8692, Fax: (403) 782-5514 Web Site: <http://www1.agric.gov.ab.ca/app21/rtw/selsubj.jsp>

Wet field conditions just prior to harvest can cause pre-harvest
sprouting in barley (*Hordeum vulgare* L.) resulting in significant
economic losses especially in barley genotypes with low seed dormancy.
Conversely, malting barley varieties with too high dormancy levels can
result in inconsistent germination, creating problems in the malt house.
Seed dormancy is defined as when viable kernels fail to germinate under
optimum conditions of moisture, oxygen, and temperature. Phenotypic
selection for sprouting resistance is challenging because the dormancy
trait is controlled by multiple genes and influenced by the environment.
Developing molecular markers linked to dormancy would be one method of
selecting for desirable levels of seed dormancy in barley. Our objective
is to identify, map and develop potential molecular markers linked to
genes affecting dormancy in 'Samson' barley. Several recombinant inbred
lines (RILs) were created by crossing 'Samson' derived lines, having
high dormancy, with hulless barley varieties 'Falcon' and 'Phoenix'.
Dormancy levels were calculated using a weighted germination index (WGI)
on the RIL population of 239 lines, originally derived from crossing
'Phoenix' and 'Samson'. This phenotyped population is currently being
analysed with SSR markers and AFLP analysis. As expected, initial
results suggest multiple QTLs throughout the barley genome, with the
most apparent marker linkages associated with seed dormancy occurring on
chromosomes: 2H, 3H, 4H, 5H, and 6H.

**Key words:** seed dormancy, marker development, *Hordeum vulgare*,
barley

## Genotypic variations in preharvest sprouting resistance and seed dormancy in barley

### Nyachiro\* J.M., J.L. Zantinge, J.H. Helm, P.E. Juskiw and D.F. Salmon

#### Field Crop Development Centre, Alberta Agriculture, Food and Rural Development, 5030 -- 50 St., Lacombe, AB T4L 1W8 Web Site: http://www1.agric.gov.ab.ca/app21/rtw/selsubj.jsp

\*Corresponding author: joseph.nyachiro@gov.ab.ca

Seed dormancy is a vital agronomic trait related to seed quality because
it determines resistance to preharvest sprouting (PHS). The aim of this
study was to evaluate if there are any genotypic differences in spike
sprouting and seed dormancy among advanced breeding lines (genotypes) of
barley and determine if there is any association between spike sprouting
and whole seed dormancy. Five separate tests comprised of 103 advanced
breeding lines and registered barley varieties were seeded in the field
in 2004 in 8-row plots of 1 x 2.5 m in three replicates arranged in a
randomized complete block design. Three intact spikes, mainly from the
primary tillers, were evaluated for resistance to sprouting resistance
in a rain simulator at 18 ^o^C. Sprouting was rated visually on a 1-5
scale (1= no visible sprouting, 5= 100% sprouted) and ratings were
converted to spike sprouting indices (SSI) that took into account the
promptness of spike sprouting. The genotypes were designated as
resistant (R) to sprouting if they had a SSI range of 3.0 to 4.0;
moderately resistant (MR) if 4.1 to 5.0; susceptible (S) if 5.1 to 6.0;
and very susceptible (VS) if \>6.0. Also whole seeds for each line were
tested for seed dormancy based on a weighted germination index (WGI).
Continuous variations were observed both in the SSI and WGI among
genotypes. There were genotypic differences in tendency for spike
sprouting ranging from 3.1 (R) to 7.3 (VS). For the SSI, the hulless
barley varieties ranged from 3.5 to 5.9, the 2-rowed ranged from 4.7 to
6.5 and the 6-rowed ranged from 3.8 to 6.5. There were wide variations
in WGI ranging between 0 (no seed germination) and 1 (100% seed
germination). For the WGI, the hulless barley varieties ranged from 0.2
to 1, the 2-rowed ranged from 0.1 to 0.9 and the 6-rowed ranged from 0
to 0.8. The cultivars Vivar and Xena (feed barley) consistently appeared
to have good resistance to spike sprouting. The correlation between the
WGI and SSI (*r* = 0.37), although significant (*P* \> 0.01), appear to
be weak.

## Number range CHAPTERUsing growing degree days to estimate maturity in small grain cereals

### Juskiw, P., Helm, J., Salmon, D., and Nyachiro, J. 

#### Field Crop Development Centre, 5030 50^th^ Street, Lacombe, AB, T4L 1W8

For over thirty years, maturities of small grain cereals have been
estimated at Field Crop Development Centre by using dry-down rates and
moisture contents at harvest. Maturity is estimated to occur at 35%
moisture content. Indicator plots of each crop type have been used each
year at each plot site to determine a linear rate of dry-down based on
Julian days. However ever year, there can be problems in getting the
indicator plots harvested during the linear phase of dry-down so we
wanted to develop a estimate for dry-down that could be used over a wide
range of environmental conditions. In previously reported work,
standardized rates of dry-down using growing degree days
(GDD=Σ\[(T~min~+T~max~)/2\] and growing season precipitation were
developed for barley, spring and winter triticale, spring and winter
wheat, and winter rye. In 2004, data were collected to determine the
validity of our GDD-based rates of dry-down. We compared GDD-based
maturities with maturities based on the indicator plot dry-down rates.
For barley, our best correlation (r=0.80) between maturities using the
two methods (n=5,181) was found when the dry-down rate included both GDD
and growing season precipitation, however the closest fit based on
similar range and mean maturities was found using a mean rate of
dry-down based solely on GDD. The 2004 data confirmed over a wide range
of environments, that using a standardized rate of dry-down based on GDD
was valid. The 2004 data will be incorporated into updated GDD and GDD
plus precipitation rates of dry-down, and a final decision will be made
on the method of determining maturities from 2005 onwards.

patricia.juskiw@gov.ab.ca

## Twelve years of barley-based rotations

### Juskiw, P., and Westling, D. 

#### Field Crop Development Centre, 5030 50^th^ Street, Lacombe, AB, T4L 1W8

In 1988 a barley-based rotation was begun at Lacombe. There were eleven
four-year rotations that were run for twelve years (three complete
cycles). After the12 years, a uniformity trial was run planting all
plots to Niska, six-row feed barley. Soil NO~3~N, PO~4~P, Na, K, and
SO~4~S were measured in the fall of 1999, with the only significant
rotation effect being found on soil nitrogen levels. When green manure
followed three years of triticale N levels were the highest, but when
green manure followed barley-canola-winter wheat levels were not
significantly elevated. A year of fallow did not elevate nitrogen
levels. Protein, acid detergent fiber, neutral detergent fiber, and
relative feed value of the biomass harvest after anthesis (about the
soft-dough stage) were not affected by rotation. Highest post-anthesis
(PA) biomass yields were found when the previous year had been fallow,
green manure or alfalfa. The lowest PA yields were following barley
(although barley the previous year was not the sole determinant of low
yields as other rotations with barley the previous year had intermediate
yields). Grain yields and test weights were not affected by rotation.
However both kernel weights and percent plumps were positively
influenced by barley-barley-canola-fallow, winter
wheat-green-manure-barley-canola, and barley-barley-alfalfa-alfalfa
rotations; while continuous barley and canola-winter wheat-peas-barley
had negative effects on these traits. Grain protein levels were highest
following continuous barley, wheat-canola-barley-barley and
triticale-triticale-triticale-green manure; and lowest following
triticale-barley-peas-barley and barley-winter rye-barley-canola.
Rotations that led to high soil N levels did not always translate into
high protein levels in the grain; and this combined with higher percent
plump and kernel weights may mean we need to rethink our recommendation
for malting barley production. Further investigation is warranted.

Patricia.Juskiw@gov.ab.ca

## Development of winter hulless barley varieties as a high value crop

### W.S. Brooks\*, C.A. Griffey and M.E. Vaughn

#### Virginia Polytechnic Institute and State University, Blacksburg, VA

Prior to the early 1990's winter barley cultivars released and grown in
the U. S. mid-Atlantic region were traditional hulled feed barley types.
Traditional hulled barley has been grown for centuries in the
mid-Atlantic region on many farms as feed for all classes of livestock.
Demand for low-fiber, high-energy grains by the vertically integrated
swine and poultry industries, and availability of brewer's distilled
grains for beef and dairy industries have resulted in greatly reduced
demand for traditional feed barley in recent years. In the mid 1990's,
the Virginia Tech Breeding Program realized that survival of winter
barley as a viable crop was dependent on development of commercially
acceptable winter hulless barley cultivars having high value traits for
specific end uses. During the past 10 years, the Virginia Tech barley
breeding program has developed hulless lines that yield 314-1129 kg
ha^-1^ higher than initial winter hulless lines developed. Many lines
have improved straw strength and grain plumpness and have better
resistance to prevalent diseases. Meanwhile, increased interest in the
use of hulless barley varieties having high energy and digestibility in
manufacturing food and fuel products, as well as feed, has accentuated
our desire to develop winter hulless barley varieties having greater
marketability in both domestic and foreign markets. Additionally, barley
grain contains health-related compounds similar to those found in oats,
therefore, adding to its appeal in the health-food sector. The use of
barley in ethanol production may soon become a reality and would provide
a viable market for hulless barley produced in the mid-Atlantic region.
We also have collaborated with nutritionists and chemists to
characterize and improve the nutritional and compositional quality of
hulless barley via breeding for specific end uses. The breeding
program's first major achievement was the release of the winter hulless
barley cultivar Doyce in 2003. In collaboration with the USDA-ARS
Eastern Regional Research Center, data on chemical and nutritional
composition, including protein, starch, lipid and beta glucan
concentration, have been obtained on most barley lines in our replicated
yield trials. To date, significant progress already has been made in the
development of winter hulless barley lines. We have developed more than
3,000 winter hulless barley populations. This year (2005), we will
advance over 350 hulless populations and evaluate 325 pure lines in
yield tests and select pure lines among nearly 9,000 hulless headrows.
Over one hundred advanced winter hulless barley lines are being
evaluated in four states (Maryland, Pennsylvania, Kentucky and
Delaware). Doyce hulless barley being produced in 2005 will be evaluated
in pilot studies for its potential use in ethanol production and as an
improved feed component in poultry rations.

\*Corresponding Author: Phone: 540 231-7624, Email: wybrooks@vt.edu

## Multiple dominant and recessive marker stock development

### Robert I. Wolfe

A set of genetic marker stocks for barley, Hordeum vulgare, have been
developed. Dominant alleles of several genes are in one doubled haploid
stock. A master recessive doubled haploid has matching recessive
alleles.

Dominant alleles in the master dominant stock with matching recessive
alleles in the master recessive: Blp Vrs1 Pre2 Zeo1 Wst7 Btr Alm Pub Kap
Hsh Srh Raw1 Rob Wax Nud Lks2.

Several more multiple recessive stocks are available. A genetic male
sterile is present in a recessive background, similar to the above
master recessive, for each of the seven chromosomes. Also incorporated
on the appropriate chromosomes are five surface wax mutants, five
dwarfs, and a few other recessive alleles.

Recessive alleles in the chromosomal stocks:

1H msg1 cer-e ert-b nec1

1H trd

2H msg2 gsh6 eog lig

3H msg5 uzu

3H als

4H msg24 glf1 lbi2 yhd(alm is not present in this stock)

5H msg19 ert-g

6H msg36 gsh4

6H cul2 dsp9

7H msg10 gsh3 brh1

These barley stocks are moderately early maturing. They were grown and
selected in western Canada, with alternate generations in growth chamber
and greenhouse.

Reference: 1996 Special Issue - Barley Genetic Newsletter. Vol. 26.

The author is grateful to Mr. Les Shugar for producing the doubled
haploid versions of the master dominant and recessive stocks.

## Genetic male sterile and xenia assisted reciprocal recurrent selection

### Robert I. Wolfe

Two pairs of barley populations were developed to illustrate the
potential for genetic male sterile and xenia assisted reciprocal
recurrent selection. They carry the following alleles:

1a) msg1, Sex1, yellow aleurone, btr1, cer‑e, vrs1, nud

1b) msg2, sex1, blue aleurone, btr1, rob, vrs1, nud

2a) msg1, Sex1, yellow aleurone, btr1, cer‑e, Vrs1, nud

2b) msg2, sex1, blue aleurone, btr1, rob, Vrs1, nud

These barley lines are spring habit, and have been selected in central
Alberta.

The idea is to speed up selection for yield and agronomic performance by
completing a full cycle of reciprocal recurrent selection per year.

Unfortunately, the blue aleurone has proven unsatisfactory for this
purpose. Its genetics are too complex, and when blue aleuroned lines are
crossed onto lines carrying the yellow aleurone, the blue aleurone does
not consistently colour up the resulting seeds.

If a practical working system is to be developed the blue aleurone
should be replaced by a gene with two easily identifiable alleles having
strong xenia penetrance. A possible candidate is Wax wax, perhaps with
the closely linked gsh3 for use as a field identifier. Wax wax can be
used in

hulled barley, whereas aleurone colour could not. Identification of waxy
versus starchy seed would be somewhat tedious, but doable. Normal versus
yellow starch might also work if it could be inserted into barley as a
single gene effect. The Sex1 sex1 gene is acceptable, along with rob as
a field identifier.

This noted, following is an explanation of the concept. There are two
generations a year, a crossing generation and a yield test. Two spring
barley populations are developed, either two‑row or six‑, and pure for
either btr1 or btr2. They must carry a different genetic male sterile,
such as msg1 and msg2. In the crossing generation, in a winter nursery,
seeds from the two populations are inter‑planted close enough to
inter‑pollinate and far enough apart to produce several tillers per
plant. Each population is normally 50% sterile and 50% heterozygous for
sterility. Only seed from the genetic male sterile plants is harvested.
The seed from each plant is identified as inter‑ or intra‑population
seed. Inter‑population seed is planted in a hill yield test in the area
for which it is to be adapted, with several seeds per hill, each hill
being from one plant. The conditions must mimic as closely as possible a
farmer\'s field for selection to be useful. The parent plants from the
winter nursery are ranked according to hill performance, and
intra‑population seed from the best ones sent south for the next
crossing nursery.

New elite germplasm can be incorporated and added to each population.

After a few cycles, F~2~ seed from the best yield test hills can be
entered into the standard breeding system in use in the program.

## Isoyield analysis of barley cultivar trials in the Canadian Prairies

### Rong-Cai Yang^1,2\*^, Daniel Stanton^2,3^, Stanford F. Blade^4^, James Helm^5^ and Dean Spaner^2^

#### ^1^ Policy Secretariat, Alberta Agriculture, Food and Rural Development, Room 300, 7000 -- 113 Street, Edmonton, AB, Canada T6H 5T6; ^2^ Dept. of Agricultural, Food and Nutritional Science, University of Alberta, Edmonton, AB, Canada T6G 2P5; ^3^ Pioneer Hi-Bred Ltd., 330 -- 127 Street S.W., Edmonton AB, Canada T6W 1A3; ^4^ International Institute of Tropical Agriculture, c/o Lambourn Ltd., Carolyn House, 26 Dingwall Road, Croydon CR9 3EE United Kingdom; ^5^Field Crop Development Centre, Alberta Agriculture, Food and Rural Development, Lacombe, AB, Canada T4L 1W8

Classification of test sites for cultivar trials into groups with
similar within-group site performance and response (isoyield groups) is
an important step towards identification of appropriate cultivars that
are best suitable for different productivity levels in farm fields. The
objective of this presentation is to determine isoyield environments in
the Canadian prairies based on the analysis of cultivar trials
consolidated from individual provinces for barley (*Hordeum vulgare*
L.). Yields for the analysis were taken from 324 replicated trials sown
at 84 sites across the prairies during 1995 -- 2003. The combined use of
regression and cluster analyses of the data normalized for averaging the
multi-year unbalanced data led to a stratification of the 84 sites into
13 isoyield groups. A comparison was made of the distributions of the
variability among and within groups according to three modes of
grouping: isoyield groups, soil zones and agroecoregions. There was more
variability among isoyield groups and correspondingly less within the
groups than that among and within soil zones and agroecoreions. Similar
contrasting pattern existed for the variance components involving
genotype-environment interaction (GEI) though the GEI variability was
generally small under all three modes of grouping. Relationships of site
sensitivity (regression coefficient) and stability (coefficient of
determination) with site productivity were shown to be a useful aid for
selecting a subset of test sites in an effort to improve efficiency and
quality of future cultivar testing. Thus, the isoyield analysis should
be a valuable tool for a meaningful subsetting of heterogeneous
environments and for a reduced GEI impact in cultivar testing and
recommendation.

\*email: rongcai.yang@gov.ab.ca

## Genetic analysis of preharvest sprouting in barley

### S.E. Ullrich\*, J.A. Clancy, H. Lee, F. Han, K. Matsui, I.A. del Blanco

#### Dept. of Crop & Soil Sciences, Washington State University, Pullman, WA 99164-6420, USA

Preharvest sprouting (PHS) can be a problem in barley production,
especially of malting barley. Rain or even very high humidity from near
physiological maturity onward can cause sprouting in spikes. This has
very serious consequences for malting grain, since rapid and complete
germination is critical. Much information has been gained by studying
the genetic control of dormancy (measured as percent germination) in
barley. The objective of this study is to determine if the germplasm
developed and QTLs discovered in previous research of dormancy can be
applied or related to the genetic control of PHS. PHS was measured in
this study as 'PHS score' based on visual sprouting in mist
chamber-treated spikes at 0 and 14 d after physiological maturity and as
'alpha-amylase activity' in kernels taken from mist chamber-treated
spikes that showed little or no visible sprouting at physiological
maturity (0 d). Germination percentage was also measured at 0 and 14
days after physiological maturity. Many QTLs for dormancy have been
previously mapped, most of which are minor and inconsistently expressed
across environments. Consistently expressed major and minor dormancy
QTLs were previously mapped to barley chromosomes 1 (7H), 4 (4H), and 7
(5H) in the U.S. Barley Genome Project Steptoe (dormant) / Morex
(non-dormant) doubled haploid mapping population. Evaluation of this
population grown in two environments (greenhouse and field) for PHS
score revealed QTL regions on all chromosomes, except chromosome 6 (6H)
and for alpha-amylase activity on all seven chromosomes from one or both
environments. However, many of the QTLs identified were minor in effect.
QTL effects for all traits analyzed ranged from 4 to 36%. The two major
dormancy QTLs previously identified on chromosome 7, as well as, the
minor ones on chromosomes 1 and 4 were confirmed in this study. Some of
the PHS score and alpha-amylase QTLs coincide with known dormancy QTLs,
but there appear to be QTLs unique to PHS, as well. Some PHS
alpha-amylase activity QTLs coincide with known malt-derived
alpha-amylase activity QTLs, but some do not. The major chromosome 7
dormancy QTLs detected from this cross are expressed during PHS, but
several previously identified minor dormancy QTLs appear to be more
important during preharvest sprouting than during after-ripening or
after-harvest dormancy conditions. Whereas, the literature frequently
equates dormancy and preharvest sprouting, it appears there is some
difference in genetic control of these two somewhat opposite traits.
Both traits are complexly inherited, but with some overlap and some
uniqueness in gene expression. In addition, several relatively major
genes seem to stand out in expression with many minor genes presumably
interacting or adding to the expression of the two traits. This study
continues with two other mapping populations previously analyzed for
dormancy. Ultimately, key QTLs will be identified, which should benefit
the breeding efforts of both six-row and two-row barley for a suitable
balance between the tendencies for preharvest sprouting and dormancy.

\* Corresponding author, e-mail address \<ullrich@wsu.edu\>

## Effects of ethylene in barley (*Hordeum vulgare* L.) tissue culture regeneration

### Jha, Ajay K.^1^, Lynn S. Dahleen^2^ and Jeff C. Suttle^2^

#### ^1^Department of Plant Sciences, North Dakota State University, Fargo, ND 58105. ^2^USDA-ARS, Fargo, ND 58105

Ethylene is a gaseous plant hormone that regulates numerous cellular
processes from germination to flowering and senescence. It is produced
under stress conditions such as tissue culture and can be
physiologically significant in-vitro due to enclosed conditions. This
study was conducted to determine genotype-dependent ethylene production
and its role in regeneration of barley (*Hordeum vulgare* L.) callus.
Six barley cultivars were examined and found to produce different
amounts of ethylene during culture. The highest regeneration was
observed in cultivars generating the most ethylene. Ethylene production
was correlated with regeneration rates (r^2^= 0.90625). There were no
significant genotype by stage interactions for either ethylene
production or green plant regeneration. The media was modified by adding
the ethylene precursor, ACC (1-amino-cyclopropane-1-carboxylic acid) or
the ethylene antagonist silver nitrate (AgNO~3~) to the media at
different stages of callus culture to determine the effects of ethylene
during plant regeneration. Highest regeneration in Morex was observed
when AgNO~3~ was added to maintenance stages (M-1, M-2) and lowest
regeneration when AgNO~3~ was applied to the regeneration stage compared
to the control. In Golden Promise, AgNO~3~ added throughout the second
maintenance and regeneration stages showed the highest regeneration
compared to control. Regeneration was significantly affected with
addition of ACC in Morex and highest when ACC was added at the second
maintenance stage. Golden Promise did not show improved regeneration
when ACC was added at any time. Regeneration was highest for the
control. Further manipulation of ethylene synthesis and/or action will
be used to identify critical timing and duration for ethylene to effects
on plant regeneration from recalcitrant genotypes. Ethylene exposure for
briefer time periods will help pinpoint the specific stages when
ethylene should be manipulated.

Corresponding author: Lynn Dahleen

e-mail: dahleenl@fargo.ars.usda.gov

## Validation of select diastatic power QTL in elite Western U. S. six-rowed spring barley germplasms

### D. Hoffman, A. Hang, and D. Obert

#### USDA-ARS Small Grains and Potato Germplasm Facility, 1691 South 2700 West Aberdeen, Idaho 83210

#### 

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Barley is a major commodity in the western US., especially when utilized
for malt production. Spring six-rowed barley germplasm adapted to the
western US. is often low in diastatic power (DP), an important malting
trait. DP quantitative trait loci (QTL) and linked markers have been
identified in a spring six-rowed mapping population grown in western
environments, but these QTL have not been validated in elite western
six-rowed backgrounds and the QTL are based on restriction fragment
length polymorphism (RFLP) markers. The objectives of this study were to
validate DP QTL in populations containing elite western germplasm and to
identify polymerase chain reaction (PCR) -based markers linked to select
DP QTL.

Low DP spring six-rowed cultivars or elite lines were crossed to
'Morex', a high DP 6-rowed barley, or to SM#42, a high DP line of the
'Steptoe'/Morex mapping population. . F~1~ plants were backcrossed to
the adapted, low DP lines two or three times, and segregating lines were
developed from the backcrossed populations. The low DP parents and
backcross-derived segregating lines were genotyped with PCR-based
markers closely aligned to five DP QTL with large effects on chromosomes
1H, 4H, and 7H. Lines homozygous for the PCR-based markers and parental
checks were planted in 4-m rows at Aberdeen, ID. Quality analysis to
determine DP levels was performed by the Cereal Crops Research Unit,
Madison, WI. Nearly all the low DP parental lines had marker alleles
that resembled either Steptoe or Morex with regard to DNA fragment size.
Two lines from Utah State University had DP-negative marker alleles for
all loci except the one on the short arm of chromosome 4H. Some had more
DP-positive alleles with a few DP-negative alleles, while others had
similar numbers of DP-positive and DP-negative alleles. Also, some
heterogeneity within lines was detected. We plan to determine if
relationships between marker genotype and DP can be identified in the
advanced populations and if other traits are affected by DP marker
selection. This study should provide useful information for the
development of six-rowed malting barleys in the western U.S.

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## The barley stem rust resistance gene product RPG1 is specifically degraded upon infection with the stem rust fungus *Puccinia graminis* f. sp. *tritici* pathotype MCC

### J. Nirmala^1^, B. Steffenson^2^ and A. Kleinhofs^1&\ 3\*^

#### ^1^ Department of Crop and Soil Sciences Washington State University, Pullman, WA-99164-USA ^2^ Department of Plant Pathology, University of Minnesota, St Paul, MN 55108, USA ^3^ School of Molecular Biosciences, Washington State University, Pullman, WA 99164, USA

Disease resistance in plants, mediated by the gene-for-gene mechanism,
involves the direct or indirect recognition by the R-gene product of
specific effector molecules produced by the pathogen. This recognition
triggers a series of signals regulating an elaborate series of defense
mechanisms by the plant. In order to understand the role of the recently
cloned barley stem rust resistance gene product RPG1 (Brueggeman et al.
2002) in resistance to the stem rust fungus *Puccinia graminis* f. sp.
*tritici*, we investigated the fate of the RPG1 protein in response to
infection with the *P. graminis* f. sp. *tritici* avirulent pathotype
MCC. Different barley lines with varying levels of resistance were
challenged with the avirulent pathotype MCC and sampled at 0, 12, 16,
20, 24 and 36h post-infection. The extracts were immuno-precipitated
with an RPG1-specific antibody. The precipitate was subjected to
SDS-PAGE, and RPG1 was visualized by western blot analysis. Though the
endogenous transcript levels of *Rpg1* remained unchanged upon infection
with the avirulent pathotype MCC (Rostoks et al. 2004), the RPG1 protein
disappeared to undetectable levels 20-24h post-infection. The
disappearance of the RPG1 protein was localized to the infected tissue
and did not spread to the adjoining leaves. The RPG1 protein was shown
to be stable in cyclohexamide translation inhibited leaves for up to 48
hrs. These results suggest that the localized disappearance of RPG1
protein is due to proteolysis, probably by the avr-gene product. Since
the mere absence of the RPG1 protein does not result in resistance, the
degradation products or the process of degradation may trigger the
signaling resulting in resistance to the stem rust infection.

^\*^ andyk@wsu.edu Tel: 1-509-335-4389

## Saturation mapping of barley chromosome 2H *Fusarium* head blight resistance QTL

### Christina Maier^1^, Deric Schmierer^1^, Thomas Drader^1^, Richard Horsley^2^, and Andris Kleinhofs^1,3^

#### ^1^Dept. of Crop and Soil Sciences, Washington State University, Pullman, WA 99164-6420, USA ^2^Dept. of Plant Sciences, North Dakota State University, Fargo, ND 58105-5051, USA ^3^School of Molecular Biosciences, Washington State University, Pullman, WA 99164-4660, USA

## 

Outbreaks of fusarium head blight (FHB), caused by *Fusarium
graminearum*, within the past two decades have caused large economic
losses to farmers in North Dakota and northwestern Minnesota due to the
reduced quality of harvested barley by blighted kernels and significant
levels of deoxynivalenol (DON), a mycotoxin produced by the fungus.
Field practices and chemical control have had limited success, putting
greater importance on a genetic approach for control. Two quantitative
trait loci (QTL) each for lower FHB severity and plant height, and one
major QTL each for DON accumulation and days to heading were mapped with
recombinant inbred lines obtained from a cross between CIho 4196, a
two-rowed resistant cultivar, and Foster, a six-rowed susceptible
cultivar (Horsley et al., submitted). These loci reside in the barley
chr. 2H region flanked by the markers ABG306 and MWG882A (bins 8-10). In
an attempt to saturate this region with markers, 29 rice chr. 4 BAC
clones with synteny to this region were blasted against the barley
expressed sequence tag (EST) database. To date, 41 of the ESTs picked by
this method have been mapped to this region. Nine other genes,
identified based on microarray analysis of the wheat-barley 2HL addition
line and comparison to Betzes and Chinese Spring controls (courtesy of
Gary Muehlbauer), were also mapped to this region on the Foster x CIho
4196 RFLP map. To date, there are a total of 26 unique loci and 67
markers in this major FHB QTL region on chr. 2. Eighteen markers have
been hybridized to the 6x cv. Morex barley BAC library, identifying 131
BAC clones as part of the physical map of the region. Three cleaved
amplified polymorphic sequences (CAPS) markers were designed for the
major FHB resistance QTL in this region, two flanking and one in the
middle, to aid in development of isolines containing fragments of this
region from CIho 4196 in a Morex cultivar background.

Christina Maier: cmaier@wsu.edu

## Map-based cloning efforts of the barley spot blotch resistance gene *Rcs5* 

### Thomas B Drader^1^, Kara A Johnson^2^, Robert S Brueggeman^1^, Hye Ran Kim^3^, Dave Kudrna^3^, Rod Wing^3^, Brian Steffeson^4^, and Andris Kleinhofs^1,\ 5^

#### 1 Crop and Soil Science, Washington State University, Pullman, WA 99164, USA 2 USDA, Washington State University, Pullman, WA 99164, USA 3 Plant Sciences Department University of Arizona, , Tucson, AZ 85721-0036, USA 4 Department of Plant Pathology, University of Minnesota, St. Paul, MN 55108-6030, USA 5 Molecular Biosciences, Washington State University, Pullman, WA 99164, USA

The *Rcs5* gene confers seedling resistance to barley spot blotch,
caused by the fungus *Cochliobolus sativus*. Spot blotch is a common and
economically important foliar disease of barley in the Midwestern United
States. Genetic mapping localized the resistance gene between the
markers MWG622 and KAJ154 on the short arm of chromosome 1(7H). A BAC
clone physical contig was constructed consisting of 4 clones, 053N3,
612G14, 452P9, and 808M17. Subclone shotgun libraries of BAC clones
612G14 and 808M17 were constructed and sequenced by the Arizona Genomics
Institute. Sequence assembly resulted in 25 contigs ranging from 1kb to
28.5kb for 808M17 and 7 contigs ranging from 0.6kb to 63.7kb for 612G14.
Analysis of the contigs\' sequence by a gene prediction program
(FGENESH) with limits within monocot genomic DNA yielded 40 putative
genes, 22 with protein homology. Of the 22 protein hits, 19 had
equivalent Triticeae EST\'s. The BAC contig overlap between 612G14 and
452P9 as well as the overlap between 452P9 and 808M17 left an
unsequenced gap of \~15kb. The BAC 452P9 38kb NotI subclone, TBD001,
covers this region and contains markers KAJ108B.2 and KAJ154, which
flank a high recombination region with 11 crossovers and potentially the
*Rcs5* gene. The subclone TBD001 sequencing is in progress. Physical and
genetic mapping of predicted genes will delimit the region of interest.
Putative candidate genes will be sequenced from resistant and
susceptible cultivars.

Corresponding author: tdrader@wsu.edu

## A gene tagging system for *Hordeum vulgare* 

### François Eudes^1\*^, André Laroche^1^, Michele Frick^1^, Jennifer Geddes^1^ and Laurian Robert^2^

#### Agriculture and Agri-Food Canada, ^1^Lethbridge Research Centre, Lethbridge, Alberta T1J 4B1, Canada and ^2^Eastern Cereal and Oilseed Research Centre, Ottawa, Ontario K1A 0C6, Canada

An efficient system for production of knockout plants for functional
analysis and gene tagging in cereals would be a significant complement
to current cereal genome analyses. *Tos17*, a rice retrotransposon
activated by tissue culture, has been successfully used for generation
of knockout plants in rice. More recently, it was reported that *Tos17*
insertions were predominately located within coding sequences and thus
provide a unique system to develop knockout plants at a relatively high
frequency. Retrotransposons have the advantage over other transposon
systems of yielding stable mutations and low copy numbers which
facilitate the identification of genes. Using a novel approach that
enables the cultivar-independent regeneration of fertile monocots,
transgenic barley T~0~ was efficiently obtained, and confirmed by
southern blot. Results suggest that *Tos17* is a very efficient system
to generate knockout plants in cereals. Under vegetative growth, the
integrated rice retrotransposon *Tos 17* is inactive based on the
absence of retrotransposase activity in barley leaves and stems. Under
tissue culture conditions, the retrotransposase activity was stimulated
and readily detected via a reverse transcription-PCR assay. A
modification in the number of *Tos17* copies in barley genomic DNA (from
callus tissues) was detected. We also regenerated fertile plants C~0~,
from 4-5 months callus culture, with novel *Tos17* insertion sites.
Using real time PCR, we report an increased copy number in C~0~ barley
genome, and new phenotype in offspring C~1~. We will discuss the
advantages of this new system for development of knockout plants in
cereals and possible impact for genomic studies in barley.

\*Corresponding author: eudesf@em.agr.ca

## Unraveling the mysteries of germination using SAGE (Serial Analysis of Gene Expression)

### Toni Pacey-Miller, Jessica White, Allison Crawford, Peter Bundock, Giovanni Cordeiro, Daniel Barbary and Robert Henry

#### Grain Foods CRC, Ltd., Centre for Plant Conservation Genetics, Southern Cross University, Lismore Australia 2480

The processes involved in malting are still somewhat a mystery on a
genetic level. SAGE (Serial Analysis of Gene Expression) is a technique
that allows rapid, detailed analysis of thousands of transcripts in a
cell. The process of SAGE relies on two principles. Firstly, a small
sequence of nucleotides from the transcript, called a "tag" can
effectively identify the original transcript from whence it came.
Secondly, linking these tags allows rapid sequencing analysis of
multiple transcripts. By examining the transcripts expressed at any time
in the cell it is possible to determine which genes and their related
proteins are being expressed at that moment in time. In this study the
gene expression profile of germinating (malting) barley is being
examined at seven intervals over a time course of 120 hours post
steeping. This will be compared to a baseline of dry ungerminated seed.
The identification of genes for improved malting quality can be
identified and examined using SAGE and ultimately used for commercial
improvement.

E-mail: tpacey@scu.edu.au