bluuebunny/crossref_metadata_2025 huggingface dataset:
MFASAT: A new alphoid DNA sequence isolated from Macaca fascicularis (Cercopithecidae, Primates)
Authors: Crovella S , Pero M Del , Marziliano N , Garagna S , Pecile V , Morgutti M , Boniotto M , Amoroso A , Montagnon D , Campa C Carlà , Lamberti L , Bigatti M P , Ardito G
Published: April 2011

A new highly repeated DNA fragment isolated from Macaca fascicularis (MFASAT) is described. Our findings obtained by sequencing, Southern blot analysis, and fluorescent in situ hybridization (FISH) on metaphasic chromosomes strongly suggest that MFASAT can be considered as a member of the alphoid DNA family characteristic of Old World monkeys. The chromosomal localization of MFASAT, obtained by FISH, showed that this alphoid DNA is present in the peri-centromeric area of all the chromosomes. MFASAT showed a high degree of conservation when compared, by sequence alignment, to other Macaca species and Papio papio as expected for species with considerable genome conservation. A low degree of homology has been found comparing M. fascicularis alphoid DNA with a more distantly related Cercopithecidae species such as Cercopithecus aethiops.Key words: alphoid DNA, Macaca fascicularis, FISH, sequence alignment.
Disease resistance gene analogs as candidates for QTLs involved in pepper-pathogen interactions
Authors: Pflieger S , Lefebvre V , Caranta C , Blattes A , Goffinet B , Palloix A
Published: April 2011

Whereas resistance genes (R-genes) governing qualitative resistance have been isolated and characterized, the biological roles of genes governing quantitative resistance (quantitave trait loci, QTLs) are still unknown. We hypothesized that genes at QTLs could share homologies with cloned R-genes. We used a PCR-based approach to isolate R-gene analogs (RGAs) with consensus primers corresponding with conserved domains of cloned R-genes: (i) the nucleotide binding site (NBS) and hydrophobic domain, and (ii) the kinase domain. PCR-amplified fragments were sequenced and mapped on a pepper intraspecific map. NBS-containing sequences of pepper, most similar to the N gene of tobacco, were classified into seven families and all mapped in a unique region covering 64 cM on the Noir chromosome. Kinase domain containing sequences and cloned R-gene homologs (Pto, Fen, Cf-2) were mapped on four different linkage groups. A QTL involved in partial resistance to cucumber mosaic virus (CMV) with an additive effect was closely linked or allelic to one NBS-type family. QTLs with epistatic effects were also detected at several RGA loci. The colocalizations between NBS-containing sequences and resistance QTLs suggest that the mechanisms of qualitative and quantitative resistance may be similar in some cases.Key words: Capsicum annuum, candidate gene, nucleotide binding site, kinase domain, quantitative trait loci.
Increased resolution of in situ hybridization signal by electron microscopy: A comparison with fluorescence microscopy
Authors: Fetni Raouf , Scott Patrick , Tihy Frédérique , Richer Claude-Lise , Lemieux Nicole
Published: April 2011

Cytogenetic studies by in situ hybridization (ISH) have proven to be valuable for gene mapping on banded chromosomes when combined with fluorescence microscopy (FISH). However, even under the best conditions, FISH technology has a resolving power inherent to light of just 0.2 µm. Its utilization is further limited by the diffusion of light coming from the fluorescent signal which covers an area considerably larger than the target DNA sequence. The development of new ISH protocols applied to electron microscopy (EMISH) should increase the resolution for cytogenetic mapping and fine chromosomal structure studies. Despite these advances, few attempts have been made which exploit this increased resolution. Here we present a detailed analysis of ISH signals obtained by fluorescence and electron microscopy methodologies to demonstrate and define the higher sensitivity obtainable by electron microscopy. This comparative study was conducted with probes of different origins: telomeric, classical satellite, alpha satellite, and single-copy DNA sequences, which provide a good reference point for later studies. We were also able to map a 200-bp cDNA probe by EMISH. This study assesses the nature of the resolution and the better definition of the EMISH signal, which confirms the greater resolution of electron microscopy as compared with that achieved with light microscopy. It also indicates that better delineation of two closely linked sequences is achieved at the electron microscopy level.Key words: In situ hybridization, electron microscopy, fluorescence microscopy, localization, repetitive and small single-copy probes.
Developing expressed sequence tags (ESTs) from polymorphic transcript-derived fragments (TDFs) in cassava (Manihot esculentaCrantz)
Authors: Suárez MC , Bernal A , Gutiérrez J , Tohme J , Fregene M
Published: April 2011

We applied the cDNA-AFLP (amplified fragment length polymorphism) technique to mRNA from the parents of a cassava (Manihot esculenta) genetic mapping population, and obtained more than 500 transcript-derived fragments (TDFs) that were unique in either parent. A subset of 50 TDFs were cloned and sequenced. Sequence alignment of the expressed sequence tags (ESTs) revealed mostly genes of unknown function. Six of the TDFs were mapped on to the cassava genetic map. We also demonstrated by genetic mapping of the TDFs, as RFLP (restriction fragment length polymorphism) markers, that TDFs are more polymorphic than random cDNAs. Generation of ESTs as differentially expressed sequences, in time or between different varieties, is proposed as a way of developing ESTs around specific traits for the candidate locus approach to mapping complex traits. Key words: cassava, genetic mapping, cDNA-AFLP technique, ESTs (expressed sequence tags).
Isolation, characterization, and chromosomal location of thetRNAMetgenes in Atlantic salmon (Salmo salar) and brown trout (Salmo trutta)
Authors: Perez J , Moran P , Garcia-Vazquez E
Published: April 2011

This work describes the isolation, characterization, and physical location of the methionine tRNA in the genome of Atlantic salmon (Salmo salar L.) and brown trout (Salmo trutta L.). An Atlantic salmon genomic library was screened using a tRNAMetprobe from Xenopus laevis. Two cosmid clones containing the Atlantic salmon tRNAMetgene were isolated, subcloned and sequenced. The tRNAMetwas mapped to metaphase chromosomes by fluorescence in situ hybridization (FISH). Chromosomal data indicated that the tDNA of methionine is tandemly repeated in a single locus in both species. Analysis of genomic DNA by Southern hybridization confirmed the tandem organization of this gene. Key words: cosmids, cloning, in situ hybridization, tRNAMet.
Genetic organization of theci-M-panregion on chromosome IV inDrosophila melanogaster
Authors: Kronhamn Jesper , Rasmuson-Lestander Åsa
Published: April 2011

The genes cubitus interruptus (ci), ribosomal protein S3A (RpS3A), and pangolin (pan) are localized within 73 kb in the cytological region 101F-102A on chromosome IV in Drosophila melanogaster. A region of 13 kb harbours the regulatory regions of both ci and pan, transcribed in opposite directions, and a 1.1-kb gene encoding RpS3A. This dense clustering gives rise to very complicated complementation patterns between different alleles in these loci. We investigated this region genetically and molecularly by use of an enhancer trap line (IA5), where the P-element was found to be inserted into the first intron of pan. Screens for imprecise excisions of the P-element were performed, and complementations between new and old established mutant lines were investigated. We found that when mutated or deleted the RpS3A gene gives rise to a Minute phenotype, and we conclude that M(4)101 encodes the ribosomal protein S3A.Key words: Drosophila melanogaster, Minute(4)101, ribosomal protein, RpS3A gene, P-element mutagenesis.
Evolution of a functionally related lactate dehydrogenase and pyruvate decarboxylase pseudogene complex in maize
Authors: Christopher Mary E , Good Allen G
Published: April 2011

A large proportion of the maize genome is repetitive DNA (60-80%) with retrotransposons contributing significantly to the repetitive DNA component. The majority of retrotransposon DNA is located in intergenic regions and is organized in a nested fashion. Analysis of an 8.2-kb segment of maize genomic DNA demonstrated the presence of three retrotransposons of different reiteration classes in addition to lactate dehydrogenase and pyruvate decarboxylase pseudogenes. Both of the pseudogenes were located within a defective retrotransposon element (LP-like element) which possessed identical long terminal repeats (LTRs) with inverted repeats at each end, a primer binding site, a polypurine tract, and generated a 5-bp target site duplication. A model describing the events leading to the formation of the LP-like element is proposed.Key words: lactate dehydrogenase, LP-like element, pseudogene, pyruvate decarboxylase, retrotransposon.
Chromosomal evidence for sibling species of the malaria vector Anopheles cruzii
Authors: Ramírez Corália CL , Dessen Eliana MB
Published: April 2011

An analysis of the ovarian polytene chromosomes of Anopheles cruzii from three localities in Southeast Brazil revealed the existence of two genetic entities within this morphologically uniform taxon. These cryptic species differed in the banding patterns of the X chromosome and 3L arm. A pattern of bands that cannot be explained by the fixation of any of the known inversions in chromosome X was revealed and named chromosomal form B to distinguish it from the standard pattern of this X chromosome, form A. Each chromosomal form is characterized by a different set of inversions. The lack of heterozygotes (A/B) for these X chromosome forms in populations where both forms coexist is evidence of absence or limited gene flow between the two groups. Key words: Anopheles cruzii, inversion polymorphism, sibling species.
Expression and genome organization of resistance gene analogs in soybean
Authors: Graham Michelle A , Marek Laura Fredrick , Lohnes David , Cregan Perry , Shoemaker Randy C
Published: April 2011

Sequence analysis of cloned plant disease-resistance genes reveals a number of conserved domains. Researchers have used these domains to amplify analogous sequences, resistance gene analogs (RGAs), from soybean and other crops. Many of these RGAs map in close proximity to known resistance genes. While this technique is useful in identifying potential disease resistance loci, identifying the functional resistance gene from a cluster of homologs requires sequence information from outside of these conserved domains. To study RGA expression and to determine the extent of their similarity to other plant resistance genes, two soybean cDNA libraries (root and epicotyl) were screened by hybridization with RGA class-specific probes. cDNAs hybridizing to RGA probes were detected in each library. Two types of cDNAs were identified. One type was full-length and contained several disease-resistance gene (R-gene) signatures. The other type contained several deletions within these signatures. Sequence analyses of the cDNA clones placed them in the Toll-Interleukin-1 receptor, nucleotide binding domain, and leucine-rich repeat family of disease-resistance genes. Using clone-specific primers from within the 3' end of the LRRs, we were able to map two cDNA clones (LM6 and MG13) to a BAC contig that is known to span a cluster of disease-resistance genes. Key words: expression, R-genes, contig, RGAs, soybean.
Relationship between pachytene synapsis, metaphase I associations, and transmission of 2B and 4B chromosomes in rye
Authors: Jiménez Guillermo , Manzanero Silvia , Puertas María J
Published: April 2011

2B rye plants selected for high (H) or low (L) B transmission rate were studied at pachytene and metaphase I of meiosis to determine the relationship between synapsis, bivalents at metaphase I, and B transmission rate. The results show that the 2 B chromosomes (Bs) form bivalents at pachytene in both the H and L lines, whereas the frequency of bivalents at metaphase I is much higher in the H than in the L line. This demonstrates that B transmission is mainly related to the proper association of Bs at metaphase I, as well as that synapsis of the 2 Bs in the L line is normal, but the bivalent is not consolidated by a chiasma in most cases. Crosses were made between 2B plants of the H and L lines in all combinations (H × H, H × L, L × H, and L × L) to obtain 4B plants. Similarly, bivalent formation at pachytene and metaphase I was studied. The results show that 4B plants of the H × H and L × L classes differ significantly at pachytene and metaphase I since the former forms more bivalents. The heterozygous 4 Bs of the H × L and L × H classes show intermediate values. The relation H × H > H × L > L × H > L × L was consistently found for the variables transmission rate, bivalents at pachytene, bivalents at metaphase I, and B mean chiasma frequency. A maternal effect was also found. Our data suggest that there are two separate mechanisms acting upon synapsis and chiasma formation in H and L B chromosomes: (i) there is variable efficiency of the control of synapsis at early stages of meiosis; and (ii) there is variable efficiency of the control of the number of chiasmata.

Key words: B chromosomes, synaptonemal complex, Secale cereale.
Molecular evolution and phylogeny of theatpB-rbcLspacer of chloroplast DNA in the true mosses
Authors: Chiang Tzen-Yuh , Schaal Barbara A
Published: April 2011

The nucleotide variation of a noncoding region between the atpB and rbcL genes of the chloroplast genome was used to estimate the phylogeny of 11 species of true mosses (subclass Bryidae). The A+T rich (82.6%) spacer sequence is conserved with 48% of bases showing no variation between the ingroup and outgroup. Rooted at liverworts, Marchantia and Bazzania, the monophyly of true mosses was supported cladistically and statistically. A nonparametric Wilcoxon Signed-Ranks test Ts statistic for testing the taxonomic congruence showed no significant differences between gene trees and organism trees as well as between parsimony trees and neighbor-joining trees. The reconstructed phylogeny based on the atpB-rbcL spacer sequences indicated the validity of the division of acrocarpous and pleurocarpous mosses. The size of the chloroplast spacer in mosses fits into an evolutionary trend of increasing spacer length from liverworts through ferns to seed plants. According to the relative rate tests, the hypothesis of a molecular clock was supported in all species except for Thuidium, which evolved relatively fast. The evolutionary rate of the chloroplast DNA spacer in mosses was estimated to be (1.12 ± 0.019) × 10-10nucleotides per site per year, which is close to the nonsynonymous substitution rates of the rbcL gene in the vascular plants. The constrained molecular evolution (total nucleotide substitutions, K approximately 0.0248) of the chloroplast DNA spacer is consistent with the slow evolution in morphological traits of mosses. Based on the calibrated evolutionary rate, the time of the divergence of true mosses was estimated to have been as early as 220 million years ago.Key words: atpB-rbcL noncoding spacer, chloroplast DNA, gene tree, molecular evolution, molecular clock, mosses, phylogeny.
The genes encoding granule-bound starch synthases at the waxy loci of the A, B, and D progenitors of common wheat
Authors: Yan Liuling , Bhave Mrinal , Fairclough Robert , Konik Christine , Rahman Sadequr , Appels Rudi
Published: April 2011

Three genes encoding granule-bound starch synthase (wx-TmA, wx-TsB, and wx-TtD) have been isolated from Triticum monococcum (AA), and Triticum speltoides (BB), by the polymerase chain reaction (PCR) approach, and from Triticum tauschii (DD), by screening a genomic DNA library. Multiple sequence alignment indicated that the wx-TmA, wx-TsB, and wx-TtD genes had the same extron and (or) intron structure as the previously reported waxy gene from barley. The lengths of the three wx-TmA, wx-TsB, and wx-TtD genes were 2834 bp, 2826 bp, and 2893 bp, respectively, each covering 31 bp in the untranslated leader and the entire coding region consisting of 11 exons and 10 introns. The three genes had identical lengths of exons, except exon1, and shared over 95% identity with each other within the exon regions. The majority of introns were significantly variable in length and sequence, differing mainly in length (1-57 bp) as a result of insertion and (or) deletion events. The deduced amino acid sequence from these three genes indicated that the mature WX-TMA, -TSB, and -TTD proteins contained the same number of amino acids, but differed in predicted molecular weight and isoelectric point (pI) due to amino acid substitutions (13-18). The predicted physical characteristics of the WX proteins matched the respective proteins in wheat very closely, but the match was not perfect. Furthermore the exon5 sequences of the wx-TmA, wx-TsB, and wx-TtD genes were different from a cDNA encoding a waxy gene of common wheat previously reported. The striking difference was that an insertion of 11 amino acids occurred in the cDNA sequence that could not be observed in the exons of the A, B, and D genes. It was noted, however, that the 3prime end of intron4 of these genes could account for the additional 11 amino acids. The sequence information from the available waxy genes identified the intron4-exon5-intron5 region as being diagnostic for sequence variation in waxy. The sequence variation in the waxy genes provides the basis for primer design to distinguish the respective genes in common wheat, and its progenitors, using PCR.

Key words: Angiosperms, Poaceae, Triticeae, Triticum monococcum, Triticium speltoides, Triticum tauschii, granule-bound starch synthase, polymerase chain reaction (PCR), molecular evolution.
Biodiversity of Asian rice gall midge (Orseolia oryzae Wood Mason) from five countries examined by AFLP analysis
Authors: Katiyar S K , Chandel G , Tan Y , Zhang Y , Huang B , Nugaliyadde L , Fernando K , Bentur J S , Inthavong S , Constantino S , Bennett J
Published: April 2011

Amplified fragment length polymorphism (AFLP) analysis was used to assess the biodiversity of one of the most important dipteran pests of cereals, the Asian rice gall midge (Orseolia oryzae Wood Mason). Larvae and pupae were collected at 15 locations in five Asian countries and preserved in 95% ethanol for storage, shipment, and DNA extraction using cetyltrimethylammonium bromide (CTAB). Although only ~1 µg of DNA was extracted from a single pupa or larva, the use of several AFLP primers in various combinations meant that this amount of DNA was sufficient to allow many DNA fingerprints to be made per individual. Fingerprints were sufficiently reproducible, especially during selective amplification, to allow the genetic diversity within a field population to be characterized. Extraction of DNA from a pool of 20 insects yielded AFLP fingerprints in which variation among individuals was sacrificed in favor of detecting differences among populations. For each location, pooled DNA was amplified with three primer pairs. A total of 261 distinct AFLP bands were identified for the 45 fingerprints. Cluster analysis, performed by the unweighted pair-group method (UPGMA), separated the populations into two distinct groups. Group I included two populations from Guangdong province of southern China and one each from Laos and Imphal in northeastern India, while group II was comprised of eleven populations from elsewhere in India (Assam, Orissa, Madhya Pradesh, Andhra Pradesh, and Kerala) and from Nepal and Sri Lanka. AFLP analysis provided insight into the origins of gall midge biotypes. In 1992, the prevailing biotype in Imphal changed from Indian biotype 3 to a new biotype 3M. Our data show that biotype 3M belongs to group I and did not arise by a recent mutation from biotype 3, which belongs to group II. By contrast, Indian biotypes 2 and 4 are likely to have diverged through recent mutation and selection, as are Chinese biotypes 1 and 4. The almost simultaneous emergence of new biotypes in Kerala and Sri Lanka during 1985-1988 was most probably coincidental, because these biotypes are not closely related. AFLP fingerprints were also able to detect sexual dimorphism in the DNA of adult gall midges and to distinguish gall midge from its major parasite Platygaster oryzae.

Key words: biotypes, Cecidomyiidae, insect, Oryza sativa, Platygaster oryzae, population, sexual dimorphism.
Construction of a long-range YAC physical map spanning the 10-cM region between the markersD18Mit109andD18Mit68on mouse proximal chromosome 18
Authors: Hsu Ssucheng J , Erickson Robert P
Published: April 2011

Four yeast artificial chromosome (YAC) contigs, physically~8 Mb, have been constructed spanning a 10-cM region on mouse proximal chromosome 18 and include the sites of 21 known genes, including those near the twirler (Tw) locus and the recently isolated Niemann-Pick type C1 (npc1) gene, formerly designated as the spm locus. This physical map consists of 49 YAC clones that cover roughly 15% of the chromosome. The physical order of 38 microsatellite sequence-tagged sites (STSs) could be assembled and confirmed based on their presence or absence in individual YACs, from proximal D18Mit109 through distal D18Mit68. These YACs provide an important resource for the further characterization and identification of known and unknown genes. The physical map has been integrated with our previously published genetic linkage map and showed an average genetic to physical distance of cM/Mb > 1.1.Key words: Mus musculus, chromosome 18, YAC contigs, physical mapping, Niemann-Pick type C1.
Varied expression of a Y-linked P[w+] insert due to imprinting inDrosophila melanogaster
Authors: Haller Bethany S , Woodruff R C
Published: April 2011

During gametogenesis, a gene can become imprinted affecting its expression in progeny. We have used the expression of a Y-linked P[w+]YAL transposable DNA element as a reporter system to investigate the effect of parental origination on the expression of the w+insert. Expression of w+was greater in male progeny when the Y chromosome, harboring the insert, was inherited from the parental male rather than from the parental female. Imprinting was not due to a genetic background influence in the males, since the only difference among the males was the parental origin of the Y chromosome. It was also observed that the genetic background can affect imprinting, since w+expression was also higher in males when the Y was derived from C(1)DX attached-X parental females rather than from C(1)RM attached-X parental females. Though the heterochromatic imprinting mechanism is unknown, a mutated Heterochromatin Protein 1 (HP1) gene, which is associated with suppression of position-effect variegation, increases expression of the w+locus in the P[w+]YAL insert, indicating that HP1 may play a role in Y chromosome packaging.

Key words: Drosophila melanogaster, heterochromatin, HP1, imprinting, P-element, Y chromosome.
Microsatellite DNA markers in Populus tremuloides
Authors: Rahman Muhammad H , Dayanandan S , Rajora Om P
Published: April 2011

Markers for eight new microsatellite DNA or simple sequence repeat (SSR) loci were developed and characterized in trembling aspen (Populus tremuloides) from a partial genomic library. Informativeness of these microsatellite DNA markers was examined by determining polymorphisms in 38 P. tremuloides individuals. Inheritance of selected markers was tested in progenies of controlled crosses. Six characterized SSR loci were of dinucleotide repeats (two perfect and four imperfect), and one each of trinucleotide and tetranucleotide repeats. The monomorphic SSR locus (PTR15) was of a compound imperfect dinucleotide repeat. The primers of one highly polymorphic SSR locus (PTR7) amplified two loci, and alleles could not be assigned to a specific locus. At the other six polymorphic loci, 25 alleles were detected in 38 P. tremuloides individuals; the number of alleles ranged from 2 to 7, with an average of 4.2 alleles per locus, and the observed heterozygosity ranged from 0.05 to 0.61, with an average of 0.36 per locus. The two perfect dinucleotide and one trinucleotide microsatellite DNA loci were the most informative. Microsatellite DNA variants of four SSR loci characterized previously followed a single-locus Mendelian inheritance pattern, whereas those of PTR7 from the present study showed a two-locus Mendelian inheritance pattern in controlled crosses. The microsatellite DNA markers developed and reported here could be used for assisting various genetic, breeding, biotechnology, genome mapping, conservation, and sustainable forest management programs in poplars.

Key words: poplar, microsatellites, genetic mapping, simple sequence repeat (SSR) markers, DNA fingerprinting.
A gene controlling sex in grapevines placed on a molecular marker-based genetic map
Authors: Dalbó M A , Ye G N , Weeden N F , Steinkellner H , Sefc K M , Reisch B I
Published: April 2011

Genetic maps of Vitis (2n = 38) have been constructed from an interspecific hybrid population of 58 seedlings of the cross 'Horizon' ('Seyval' × 'Schuyler') × Illinois 547-1 (V. cinerea B9 × V. rupestris B38). The maps were initially constructed based on 277 RAPD (random amplified polymorphic DNA) markers using a double-pseudotestcross strategy. Subsequently, 25 microsatellites, 4 CAPS (cleaved amplified polymorphic sequence), and 12 AFLP (amplified fragment length polymorphism) markers were added to the maps. Another 120 markers, mostly those segregating 3:1, were also assigned but not positioned on the linkage groups in the two maps. The 'Horizon' map consisted of 153 markers covering 1199 cM, with an average map distance of 7.6 cM between markers. The Illinois 547-1 map had 179 markers covering 1470 cM, with an average map distance of 8.1 cM. There were 20 linkage groups in each map, one more than the basic number of chromosomes in grapes. Ten linkage groups in each map were identified as homologous using 16 microsatellite and 2 CAPS markers polymorphic in both parents. A single locus controlling sex in grapes mapped close to a microsatellite marker. These maps provide enough coverage of the genome for QTL (quantitative trait loci) analysis and as a starting point for positional gene cloning in grapes.

Key words: Vitis, RAPD, microsatellite, SSR, CAPS.
Sequence analysis of trinucleotide repeat microsatellites from an enrichment library of the equine genome
Authors: Tozaki T , Inoue S , Mashima S , Ohta M , Miura N , Tomita M
Published: April 2011

Microsatellites are useful tools for the construction of a linkage map and parentage testing of equines, but only a limited number of equine microsatellites have been elucidated. Thus, we constructed the equine genomic library enriched for DNA fragments containing (CAG)n repeats. The enriched method includes hybridization-capture of repeat regions using biotin-conjugated oligonucleotides, nucleotide substrate-biased polymerase reaction with the oligonucleotides and subsequent PCR amplification, because these procedures are useful for the cloning of less abundant trinucleotide microsatellites. Microsatellites containing (CAG)n repeats were obtained at the ratio of one per 3-4 clones, indicating an enrichment value about 104-fold, resulting in less time consumption and less cost for cloning. In this study, 66 different microsatellites, (CAG)n repeats, were identified. The number of complete simple CAG repeats in our clones ranged 4-33, with an average repeat length of 8.8 units. The microsatellites were useful as sequence-tagged site (STS) markers. In addition, some clones containing (CAG)n repeats showed homology to human (CAG)n-containing genes, which have been previously mapped. These results indicate that the clones might be a useful tool for chromosome comparison between equines and humans.

Key words: microsatellites, (CAG)n repeats, equine.
Flocculation in methanogens, a comparative study of Methanosarcina barken strains Jülich and Fusaro
Authors: Scherer P. A. , Bochem H. -P. , Davis J. D. , White D. C.
Published: April 2011

Two strains of Methanosarcina barkeri grown on methanol as substrate were investigated for their ability to aggregate: the new, flocculent strain Jülich formed stable flocs of several millimetres in diameter during rapid growth on a methanol-containing medium ("bread-crumb growth"). When observed with an electron microscope, the Jülich strain showed a unique parenchymatic texture with thick-walled cells inside a floc, and coccoidlike cells on the periphery. In contrast to the Jülich strain, the strain Fusaro grew on methanol in dispersed form, and generated macroscopically visible clumps only under poor growth conditions, induced, for example, by calcium deficiency. The formation of large cell aggregations of the Fusaro strain could also be induced during growth in the presence of 0.01% of the stain Calcofluor which is known to interact specifically with β-1,4 and β-1,3 glucan moieties. Sugar analyses revealed a different pattern for both strains: the exopolymer of the flocculent strain Jülich contained half the rhamnose, a third the fucose, equal amounts of mannose and glucose, but four times more glucuronic acid and arabinose than the strain Fusaro grown in the dispersed form. The high proportion of glucuronic acid in the exopolymer of the flocculating form is of particular interest because of the role of uronic acids in adhesion.
Trypanosoma musculi infection in the CBA/N mouse
Authors: Vigeant P. , Dubois R. , Viens P.
Published: April 2011

The infection of B-cell deficient CBA/N mice by Trypanosoma musculi resulted in a higher parasitaemia, delayed recovery, and a defective antibody response (IgM and IgG1). CBA/N mice were thus less efficient than CBA/J in controlling the infection. These results indicate that B-cells are involved in the immunological control of this parasite.
Competition for nodulation of field-grown soybeans by strains of Rhizobium fredii
Authors: McLoughlin Thomas J. , Alt Scott G. , Owens P. Ann , Fetherston Corrine
Published: April 2011

Nodulation of Glycine max (L) Merr. by six Rhizobium fredii strains was measured in two Midwestern fields containing high indigenous populations of Bradyrhizobium japonicum (3 × 105/gm soil). The soils were inoculated with antibiotic-resistant mutants using liquid inoculum at two levels on soybean cv. Peking and cv. Jacques 130. Strain establishment was measured 40 days after planting. In the first year, USDA206, USDA217, and USDA257 were the most competitive strains, occupying greater than 50% of the nodules on cv. Peking in both soils. None of the strains were competitive on Jacques 130. In the second growing season, all nodules were formed by the indigenous population on both cultivars, suggesting that these fast-growing strains do not persist in Midwestern soils.
DISLOCATION DYNAMICS IN DEFORMATION AND RECOVERY
Authors: Li James C. M.
Published: April 2011

Phenomenological equations between dislocation velocity, effective shear stress, temperature, and pressure are presented. It is suggested that the term "activation volume" be properly assigned to the pressure dependence of dislocation velocity. Indirect methods of determining velocity–stress relations are reviewed, and a new method is introduced using the stress–time data in stress relaxation. Some results are reported for high-purity iron, LiF, and NaCl crystals. Phenomenological relations near absolute zero are examined in the light of the third law of thermodynamics. These relations are illustrated with the deformation behavior of potassium at liquid helium temperatures.
MICROPLASTICITY DETECTED BY AN ACOUSTIC TECHNIQUE
Authors: Fisher R. M. , Lally J. S.
Published: April 2011

Evidence for the formation of slip lines and slip bands has usually been obtained from slip-line replica and etch-pit techniques. An apparatus has been built to detect the formation of small slip events within a crystal by monitoring small longitudinal oscillations of the test specimen which occur when the load is momentarily relaxed. Each load drop results in a burst of acoustic energy, some of which are detected and recorded electronically. Results are presented of measurements of the frequency and magnitude of acoustic pulses from single-crystal specimens during plastic flow. They suggest that slip events occur cooperatively and that dislocation velocities during yielding may be as high as 103 cm/sec.
Stoichiometry control mechanisms for bias-sputtered zinc-oxide thin films
Authors: Brett M. J. , Parsons R. R.
Published: April 2011

We have deposited transparent, conducting ZnO thin films of resistivity 3 × 10−3 Ω∙cm by the technique of DC planar-magnetron sputtering in a reactive Ar–O2 atmosphere, incorporating a reactive gas baffle and substrate RF discharge. The substrate discharge was found to increase the oxygen content of the growing film. Films at low values of the RF-induced substrate self-bias voltage were characterized by a brown colour, resistivities of about 4 × 10−2 Ω∙cm, and composition ZnO0.8. Films at higher bias voltages of −80 V were clear with resistivities of 3 × 10−3 Ω∙cm and a composition approaching stoichiometric ZnO. The oxidation of the films by the RF discharge was shown to occur through preferential resputtering and re-evaporation of excess zinc and by activation and ion-plating of oxygen species. Resputtering and re-evaporation rates were found to be enhanced above that expected for bulk Zn, owing to the loosely bound nature of surface adatoms during film growth.
Shuttle vectors for genetic manipulations inUstilago maydis
Authors: Kojic Milorad , Holloman William K
Published: April 2011

Shuttle vectors with new or improved features were constructed to enable facile genetic manipulations in the plant pathogen Ustilago maydis. Sets of plasmids selectable in media containing geneticin, carboxin, nourseothricin, or hygromycin, able to replicate autonomously, to transform U. maydis by integration, and to express foreign genes under control of the homologous glyceraldehyde-3-phosphate dehydrogenase promoter, were built upon a common pUC19 vector backbone. This permits a large number of choices for a cloning site, blue/white screening for recombinant plasmids, rapid transfer of a cloned DNA fragment between plasmids, and choice of several dominant drug-resistance markers for selection in U. maydis.Key words: G418, carboxin, nourseothricin, hygromycin, expression vectors.
Effects of non-ionic surfactants on the uptake and hydrolysis of fluoresceindiacetate by alkane-oxidizing bacteria
Authors: Bruheim Per , Eimhjellen Kjell
Published: April 2011

Biological effects of non-ionic surfactants on alkane-oxidizing bacteria were studied by assessing their influence on the uptake of prefluorochrome fluoresceindiacetate (FDA) and its intracellular hydrolysis to fluorescein. Both decreasing and increasing rates of hydrolysis as a consequence of the presence of surfactants were observed. The surfactants influenced the uptake of FDA, but not its intracellular hydrolysis. The effects of the surfactants on the uptake rate depended strongly on the structure and physico-chemical properties of the surfactants. There was no qualitative or significant quantitative difference in surfactant susceptibility between induced (alkane grown) and non-induced bacteria (acetate grown), even though the induced cells possess greater cell surface hydrophobicity.Key words: fluoresceindiacetate, bacteria, surfactants, alkane.
Characterization of subterranean bacteria in the Hungarian Upper Permian Siltstone (Aleurolite) Formation
Authors: Farkas Gyöngyi , Gazsó L G , Diósi G
Published: April 2011

The main purpose of this work was to study the microbiology of the Hungarian Upper Permian Siltstone (Aleurolite) Formation, to assess the safety of future underground repositories for nuclear waste. Sixty-seven air, groundwater, technical water, rock, and surface samples were collected aseptically from different depths. The number of aerobic and anaerobic isolates was 277. The mesophilic minimum and maximum CFU counts of the air samples were 1.07-5.84 × 102·mL-1 (aerobic) and 0.22-1.04 × 102·mL-1 (anaerobic), respectively; those of the water samples were 0.39-1.25 × 105·mL-1 (aerobic) and 0.36-3.9 × 103·mL-1 (anaerobic); those of the technical water samples were 0.27-5.03 × 106·mL-1 (aerobic) and 4 × 105 - >106·mL-1 (anaerobic); and those of the aleurolite samples were 2.32 × 102 - 2.47 × 105·g-1 (aerobic) and 0.45-9.5 × 102·g-1 (anaerobic). In the groundwater, the thermophilic aerobic bacteria count was 0-2.4 × 102·mL-1 and the thermophilic anaerobic bacteria count was 0.43-4.6 × 104·mL-1. The gases produced by the 16 gas-forming isolates were CO2 (aerobic isolates), and CO2 and H2 (anaerobic isolates). About 20% of the aerobic isolates produced siderophores. The proportions of organic acid producers were lowest in aerobic and anaerobic isolates from the aleurolite, 13% and 14%, respectively. The highest proportions of acid producers in the aerobic and anaerobic isolates from the air samples were 63% and 54%. Altogether 160 of the aerobic isolates and 52 of the anaerobic isolates were spore formers. The radiosensitivity of the aerobic isolates was also determined; the D10 values of the sporeformers ranged between 0.8-2.44 kGy. Our results indicate that the sulfate-reducing bacteria and the production of complexing agents (siderophores) may contribute to the mobilization of radionuclides from underground repositories. As well, microbial gas production can influence the environmental conditions. The variability in bacterial radiotolerance indicates the biodiversity at this potential disposal site. These facts must be considered during the planning of a nuclear waste repository.Key words: nuclear waste disposal, microbial gas production, siltstone formation, siderophore production, radiosensitivity.
Optimizing the expression of a monoclonal antibody fragment under the transcriptional control of the Escherichia coli lac promoter
Authors: Donovan Robert S , Robinson Campbell W , Glick Bernard R
Published: April 2011

The expression of a monoclonal antibody Fab fragment in Escherichia coli strain RB791/pComb3, induced with either lactose or isopropyl-beta-D-thiogalactoside (IPTG), was compared to determine if lactose might provide an inexpensive alternative to induction with IPTG. Induction of Fab expression imposed a metabolic load on the recombinant cells, resulting in lower final cell yields compared to the non-induced controls. An IPTG concentration of 0.05 mM was sufficient to achieve maximal expression of soluble Fab protein when inducing in the early-, mid-, or late-log phases of batch cultures grown using either glucose or glycerol as a carbon source. The largest overall yield of Fab fragments when using 0.05 mM IPTG was achieved by increasing the final yield of cells through glycerol feeding following induction in late-log phase. Lactose was as effective as IPTG for inducing Fab expression in E. coli RB791/pComb3. The greatest overall level of Fab expression was found when cells grown on glycerol were induced with 2 g/L lactose in late-log phase. Since the cost of 0.05 mM of IPTG is significantly greater than the cost of 2 g/L lactose, lactose provides an inexpensive alternative to IPTG for inducing the expression of Fab fragments, and possibly other recombinant proteins, from the E. coli lac promoter.
Fatty-acid biosynthesis in a branched-chain α-keto acid dehydrogenase mutant ofStreptomyces avermitilis
Authors: Cropp T Ashton , Smogowicz Adam A , Hafner Edmund W , Denoya Claudio D , McArthur Hamish AI , Reynolds Kevin A
Published: April 2011

Fatty-acid biosynthesis by a branched-chain alpha-keto acid dehydrogenase (bkd) mutant of Streptomyces avermitilis was analyzed. This mutant is unable to produce the appropriate precursors of branched-chain fatty acid (BCFA) biosynthesis, but unlike the comparable Bacillus subtilis mutant, was shown not to have an obligate growth requirement for these precursors. The bkd mutant produced only straight-chain fatty acids (SCFAs) with membrane fluidity provided entirely by unsaturated fatty acids (UFAs), the levels of which increased dramatically compared to the wild-type strain. The levels of UFAs increased in both the wild-type and bkd mutant strains as the growth temperature was lowered from 37°C to 24°C, suggesting that a regulatory mechanism exists to alter the proportion of UFAs in response either to a loss of BCFA biosynthesis, or a decreased growth temperature. No evidence of a regulatory mechanism for BCFAs was observed, as the types of these fatty acids, which contribute significantly to membrane fluidity, did not alter when the wild-type S. avermitilis was grown at different temperatures. The principal UFA produced by S. avermitilis was shown to be delta9-hexadecenoate, the same fatty acid produced by Escherichia coli. This observation, and the inability of S. avermitilis to convert exogenous labeled palmitate to the corresponding UFA, was shown to be consistent with an anaerobic pathway for UFA biosynthesis. Incorporation studies with theS. avermitilis bkd mutant demonstrated that the fatty acid synthase has a remarkably broad substrate specificity and is able to process a wide range of exogenous branched chain carboxylic acids into unusual BCFAs.Key words: Streptomyces avermitilis, fatty acid biosynthesis, avermectin.
Isolation of a new laccase isoform from the white-rot fungiPycnoporus cinnabarinusstrain ss3
Authors: Otterbein Ludovic , Record Eric , Chereau David , Herpoël Isabelle , Asther Marcel , Moukha Serge M
Published: April 2011

Two extracellular laccase isoforms (Lac I and Lac II) produced by the white-rot fungus Pycnoporus cinnabarinus from the monokaryotic strain ss3 were purified from ferulic-acid-induced liquid culture medium using ammonium sulphate precipitation, followed by anion-exchange chromatography on a Mono Q column. Strain ss3 is the first generation of the parental strain P. cinnabarinus I-937. The new isolated isoform, Lac II, consists of an 86 000 molecular weight protein as determined by SDS polyacrylamide gel electrophoresis. The N-terminal amino acid sequences of both isoforms were determined, and compared to known laccase protein sequences of other organisms.Key words: oxydo-reductase, filamentous fungi, purification.
Survival of Escherichia coli in freshwater: β-D-Glucuronidase activity measurements and characterization of cellular states
Authors: Petit Muriel , George Isabelle , Servais Pierre
Published: April 2011

A rapid enzyme assay measuring β-D-glucuronidase activity of Escherichia coli was tested in survival experiments after discharge of E. coli in river water. Enzyme activity was compared with several analyses performed to characterize cellular states under stressful conditions. Enzyme activity remained stable under starvation and light stress conditions despite losses of culturability, respiratory activity, and cytoplasmic membrane integrity. β-D-Glucuronidase activity followed the pattern of genetic and morphologic cell integrity. The tested enzyme assay seems well adapted to study the fate of fecal coliforms in survival experiments, and appears to be a rapid and efficient way to estimate the microbiological quality of surface waters.Key words: β-D-glucuronidase activity, Escherichia coli, survival, freshwater, cellular states.
Molecular cloning, heterologous expression, and functional characterisation of a malate synthase gene fromStreptomyces coelicolorA3(2)
Authors: Loke Paxton , Sim Tiow-Suan
Published: April 2011

With the rapid generation of genetic information from the Streptomyces coelicolor genome project, deciphering the relevant gene products is critical for understanding the genetics of this model streptomycete. A putative malate synthase gene (aceB) from S. coelicolor A3(2) was identified by homology-based analysis, cloned by polymerase chain reaction, and fully sequenced on both strands. The putative malate synthase from S. coelicolor has an amino acid identity of 77% with the malate synthase of S. clavuligerus, and possesses an open reading frame which codes for a protein of 540 amino acids. In order to establish the identity of this gene, the putative aceB clones were subcloned into the expression vector pET24a, and heterologously expressed in Escherichia coli BL21(DE3). Soluble cell-free extracts containing the recombinant putative malate synthase exhibited a specific activity of 1623 (nmol·mg-1·min-1), which is an increment of 92-fold compared to the non-recombinant control. Thus, the gene product was confirmed to be a malate synthase. Interestingly, the specific activity of S. coelicolor malate synthase was found to be almost 8-fold higher than the specific activity of S. clavuligerus malate synthase under similar expression conditions. Furthermore, the genomic organisation of the three Streptomyces aceB genes cloned thus far is different from that of other bacterial malate synthases, and warrants further investigation.Key words: primary metabolism, polymerase chain reaction, glyoxylate pathway.
Effects of theFusariumspp. mycotoxins fusaric acid and deoxynivalenol on the growth ofRuminococcus albusandMethanobrevibacter ruminantium
Authors: May Harold D , Wu Qingzhong , Blake Cheryl K
Published: April 2011

The Fusarium spp. mycotoxins fusaric acid and deoxynivalenol (DON) were tested for antimicrobial activity against Ruminococcus albus and Methanobrevibacter ruminantium. The growth of both organisms was inhibited by fusaric acid as low as 15 µg/mL (84 µM) but not by DON, at levels as high as 100 µg/mL (338 µM). No synergistic inhibitory effect was observed with DON plus fusaric acid. Neither organism was able to adapt to the fusaric acid and responses of each organism to the compound were different. The optical density (OD) maximum for R. albus, but not for M. ruminantium, was diminished after 28 days incubation at concentrations of fusaric acid below 240 µg/mL. Inhibition of R. albus started before significant growth had occurred, while M. ruminantium doubled twice before the onset of inhibition. Responses to picolinic acid, an analog of fusaric acid, were also dramatically different between the two microorganisms with M. ruminantium exhibiting a severe lag followed by a complete recovery of growth, while R. albus was only slightly inhibited with no lag. These results suggest that the mechanism of fusaric acid inhibition is specific to each microorganism. This is the first demonstration of the common mycotoxin fusaric acid inhibiting the growth of rumen bacteria.Key words: mycotoxins, fusaric acid, deoxynivalenol, Ruminococcus albus, Methanobrevibacter ruminantium.
Evaluation of polyclonal-antibody-based immunoassays for detection of Sclerotinia sclerotiorum on canola petals, and prediction of stem rot
Authors: Bom M , Boland G J
Published: April 2011

The potential for using polyclonal-antibody-based immunoassays for detection of Sclerotinia sclerotiorum on canola petals as part of a disease prediction model was investigated. A commercial ELISA kit designed for Sclerotinia homoeocarpa was evaluated for specificity to S. sclerotiorum in comparison to other Sclerotinia spp., and known phyllosphere fungi. This polyclonal-antibody-based kit cross-reacted with antigens from other Sclerotinia spp., and fungi, and absorbance values obtained from S. sclerotiorum-infested canola petals were poorly correlated with percentages of infested petals as determined by plating on semi-selective medium, except when petals were incubated at high humidity for 24 h at 20°C-22°C prior to ELISA evaluation. Additonal polyclonal antibodies were prepared from mycelial and semi-purified cell wall antigens, and these antibodies were more specific to S. sclerotiorum than the ELISA kit. However, absorbance values obtained from S. sclerotiorum-infested canola petals were poorly correlated with percentages of infested petals as determined by plating on semi-selective medium. The results are discussed in relation to the use of polyclonal-antibody-based immunoassays for the prediction of epidemics or crop risk from sclerotinia stem rot of canola.Key words: sclerotinia stem rot of canola, Brassica rapa, Brassica napus, enzyme-linked immunosorbent assay, ELISA, serological detection, immunoassay.