Plant Breeding Science

妻⿅良亮（mega@yamaguchi-u.ac.jp）

植物育種学研究分野では、植物に普遍的に存在する⼀次代謝物から植物特有の⼆次代謝物のプロファイルを基に、植物の形質特異的な代謝物(バイオマーカー)の合成経路を特定し、その関連遺伝⼦を⽬標に育種を⾏うことで、耐乾性や節⽔性に優れた植物を作出することを最終⽬標として研究を⾏っています。

7YLvPH

L6YP
Ta
L9YPHv
L5
TaPY

ManDry形質評価および系統選抜
バイオマーカー探索• 乾燥ストレス耐性

PYL4

PYL 5

PYL
6
PY
L13PYL11• 節⽔性Root

TaPYL1TaPYL2TaPYL3TaPYL4TaPYL5TaPYL6TaPYL7TaPYL8TaPYL9乾燥ストレス下での種⼦の形態解析

各種サンプル間⽐較
Agilent 6420
Triple Quadrupole LC/MSryIm speb e
Im ibe db d
Yo ibe seeu d d
Yo ng see 24
u le d h
Ad ng af 48u sh
Ad lt le temu aFl lt s fag te
Be le mfo af
Af rete flo
M r fl w
at ow erur e ine rin gsp giked

D

植物体の形態⽐較

Ta L7
PY
Ta L8
PYL9100HighTaPYL1TaPYL2TaPYL3TaPYL4灌漑区、⾮灌漑区での⽣育⽐較
TaPYL5TaPYL6TaPYL7TaPYL8TaPYL9PY6hc c

代謝物反応経路解析

Low

原因遺伝⼦の同定
TaPP2C680cAOSd6h300b602000hLC-MSあるいはGC-MSなどの分析

c0hbc

次世代シークェンサーを利⽤した遺伝⼦

40a20a0a100aab0abcaab

3,016

遺伝⼦発現による、バイオマーカー合成に関わる遺伝⼦の同定

ull
L8L17L21Null
L8L17L21N

1L21Null
L8L17lL8L17ulN

1,065

機器を⽤いたメタボローム解析

ab

発現あるいはゲノム解析
L26NaClTaPYL1TaPYL2TaPYL3TaPYL4TaPYL5TaPYL6TaPYL7TaPYL8TaPYL92L1
PY

TaABA

c 6 12 24 6 12 24 6 12 24 6 12 24 6 12 24

ABA

2WaterShootPYL7Ta

研究テーマ
１．植物⽣理学、分⼦⽣物学的解析による形質の評価
NATURE PLANTS
２．メタボローム解析による形質特異的バイオマーカーの探索
c３．新しい形質を持った作物の開発

612813

ゲノム解析による、バイオマーカー合成に関わる遺伝⼦の同定

of the expected insertion site (Fig. 2A). Homozygous T-DNA
mutants were compared with wild-type plants, and disruption ofneither gene resulted in obvious morphological defects. Metabolite
profiles comparing levels of M217 showed that accumulation ofM217 was severely decreased in daar1 but unaffected in daar2
compared with wild type (Fig. 2B). These data indicate thatDAAR1 is responsible for the majority of M217 accumulation.

実験系統の作出
組換え⾃殖系統(RIL)など交配によるIdentification of N-Malonyl-D-allo-Isoleucine. The chemical identityof M217 was needed to further elucidate biochemical basis forthe genetic link found between this metabolite and DAAR1. Byusing a variety of chemical analysis techniques, including MS,MS/MS, NMR, and HPLC, we determined that M217 is NMDIle (Fig. 3).
The accurate mass and isotope profile information obtained
Ref: Strauch RC et al. (2015)
from the MS data suggested a molecular formula of C9H15NO5.
–10–50510–10–505100More structural insights were obtained by NMR analysis on M217
log
[foldchange]
log2[fold change]
purified from Arabidopsis leaf tissue using semipreparative
Fig. 1. Manhattan 2plot showing GWAS results for M172T666. The −log10
HPLC. All assignments were done using 1H, 13C, heteronuclear
P values for association with M172T666 for each SNP (y axis) are plottedagainst chromosome positions (x axis). The horizontal dashed line corresingle quantum coherence (HSQC), and correlation spectroscopy
spondstoa
Bonferroni-correctedα<0.05.
(COSY) spectra (Table 1). The COSY spectrum was especiallya, Phylogenetic tree for ABA receptors (black, Arabidopsis thaliana; red, Triticum aestivum
useful in identifying the isoleucine moiety as there is a high dethe maximum-likelihood method in MEGA 6.06 followed by bootstrapping with 1,000
gree of proton–proton coupling (J-coupling) in the R-group.
Except for DAP epimerase, which is required for the biosynthesis
Once
the R-group was assigned, comparison with spectra of!=!3 biologically independent samples;
centralvaluesandProRerrorbars represent!s.d.).
of Lysin bacteria
and plants,
in Trypanosoma
cruzi, themeans!±
proisoleucine standards confirmed the identity of the amino acid.
tozoan parasite responsible for Chagas disease, has been the onlyof the malonyl derivatization was based onnder various stresses (c) and at different
developmental stages (d). Water was used asIdentificationaknown eukaryotic PLP-independent racemase (15). The Arabidopsis
several lines of chemical evidence. Two carbonyl groups and aamino acidracemasethat wemannitol
describe in (Man)this paperoris dehydrationa news were subjected to 20!µM ABA, 150!mM
NaCl,200!mM
member of the PLP-independent family, making its identification a methylene (CH2) group remained to be accounted for in theNMRevels were evaluated according to copy
numberandnormalized
relative to TaActin (n!=!3 spectrum after isoleucine was assigned. Although no addisignificant
additionto thisclass of enzymes.
tional direct structural information could be obtained from NMR,
ponsive genes in Null and TaPYLox seedlings.
Seven-day-old seedlings were treated the lower-than-expected integration of C8 methylene (CH2) in theResults
1H spectrum was suggestive of a β-dicarbonyl group. Methylene
GenomeWideAssociationAnalysis
RevealedaStrong
QTLforanndependent samples; central values and bars represent means!±!s.d.). Different letters
protons in this group exchange with the solvent through a ketoUnknown Metabolite. A total of 440 natural A. thaliana acces-

実験系統の作出

新規系統の開発

育成系統への原因遺伝⼦の戻し交配もしくはゲノム編集など

⼈⼯気象機を利⽤した実験
系統の促成栽培