A rice gene activation/knockout mutant resource for high throughput functional genomics.
Using transfer DNA (T-DNA) with functions of gene trap and gene knockout and activation tagging, a mutant population containing 55,000 lines was generated. Approximately 81% of this population carries 1-2 T-DNA copies per line, and the retrotransposon Tos17 was mostly inactive in this population during tissue culture. A total of 11,992 flanking sequence tags (FSTs) have been obtained and assigned to the rice genome. T-DNA was preferentially ( approximately 80%) integrated into genic regions. A total of 19,000 FSTs pooled from this and another T-DNA tagged population were analyzed and compared with 18,000 FSTs from a Tos17 tagged population. There was difference in preference for integrations into genic, coding, and flanking regions, as well as repetitive sequences and centromeric regions, between T-DNA and Tos17; however, T-DNA integration was more evenly distributed in the rice genome than Tos17. Our T-DNA contains an enhancer octamer next to the left border, expression of genes within genetics distances of 12.5 kb was enhanced. For example, the normal height of a severe dwarf mutant, with its gibberellin 2-oxidase (GA2ox) gene being activated by T-DNA, was restored upon GA treatment, indicating GA2ox was one of the key enzymes regulating the endogenous level of GA. Our T-DNA also contains a promoterless GUS gene next to the right border. GUS activity screening facilitated identification of genes responsive to various stresses and those regulated temporally and spatially in large scale with high frequency. Our mutant population offers a highly valuable resource for high throughput rice functional analyses using both forward and reverse genetic approaches.
Hsing
Yue-Ie
YI
Institute of Plant and Microbial Biology, Academia Sinica, Nankang, Taipei 115, Taiwan, ROC.
Chern
Chyr-Guan
CG
Fan
Ming-Jen
MJ
Lu
Po-Chang
PC
Chen
Ku-Ting
KT
Lo
Shuen-Fang
SF
Sun
Peng-Kai
PK
Ho
Shin-Lon
SL
Lee
Kuo-Wei
KW
Wang
Yi-Chieh
YC
Huang
Wen-Lii
WL
Ko
Swee-Suak
SS
Chen
Shu
S
Chen
Jyh-Long
JL
Chung
Chun-I
CI
Lin
Yao-Cheng
YC
Hour
Ai-Ling
AL
Wang
Yet-Walt
YW
Chang
Ya-Chi
YC
Tsai
Min-Wei
MW
Lin
Yi-Show
YS
Chen
Yin-Chin
YC
Yen
Hsing-Mu
HM
Li
Charng-Pei
CP
Wey
Chiu-Kai
CK
Tseng
Ching-Shan
CS
Lai
Ming-Hsing
MH
Huang
Sheng-Chung
SC
Chen
Liang-Jwu
LJ
Yu
Su-May
SM
D001483:Base Sequence; D004269:DNA, Bacterial / Q000235:genetics; D018744:DNA, Plant / Q000235:genetics; D018506:Gene Expression Regulation, Plant; D017343:Genes, Plant; D005822:Genetic Vectors; D023281:Genomics; D009154:Mutation; D012275:Oryza / Q000235:genetics*; D030821:Plants, Genetically Modified; D016324:Sequence Tagged Sites; D015533:Transcriptional Activation
D004269:DNA, Bacterial; D018744:DNA, Plant; C036483:T-DNA