Stable transformation of rice (Oryza sativa L.) via microprojectile bombardment of highly regenerative, green tissues derived from mature seed.
A highly efficient and reproducible transformation system for rice ( Oryza sativa L. cv. Taipei 309) was developed using microprojectile bombardment of highly regenerative, green tissues. These tissues were induced from mature seeds on NB-based medium containing 2,4-dichlorophenoxyacetic acid (2,4-D), 6-benzylaminopurine (BAP) and high concentrations of cupric sulfate under dim light conditions; germinating shoots and roots were completely removed. Highly regenerative, green tissues were proliferated on the same medium and used as transformation targets. From 431 explants bombarded with transgenes [i.e. a hygromycin phosphotransferase ( hpt) gene plus one of a wheat thioredoxin h ( wtrxh), a barley NADP-thioredoxin reductase ( bntr), a maize Mutator transposable element ( mudrB) or beta-glucuronidase ( uidA; gus) gene], 28 independent transgenic events were obtained after an 8- to 12-week selection period, giving a 6.5% transformation frequency. Of the 28 independent events, 17 (61%) were regenerable. Co-transformation of the second introduced transgene was detected in 81% of the transgenic lines tested. Stable integration and expression of the foreign genes in T(0) plants and T(1) progeny were confirmed by DNA hybridization, western blot analyses and germination tests.
Cho
M-J
MJ
Department of Plant and Microbial Biology, University of California, Berkeley 94720, USA. mjcho@nature.berkeley.edu
Yano
H
H
Okamoto
D
D
Kim
H-K
HK
Jung
H-R
HR
Newcomb
K
K
Le
V K
VK
Yoo
H S
HS
Langham
R
R
Buchanan
B B
BB
Lemaux
P G
PG
D001483:Base Sequence; D019470:Biolistics; D002934:Cinnamates / Q000494:pharmacology; D004274:DNA, Recombinant / Q000235:genetics; D004351:Drug Resistance / Q000235:genetics; D015870:Gene Expression; D017343:Genes, Plant; D006921:Hygromycin B / Q000031:analogs & derivatives* / Q000494:pharmacology; D012275:Oryza / Q000187:drug effects / Q000235:genetics* / Q000254:growth & development / Q000502:physiology; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000235:genetics; D030821:Plants, Genetically Modified; D012038:Regeneration; D014170:Transformation, Genetic
D002934:Cinnamates; D004274:DNA, Recombinant; D006921:Hygromycin B; C026273:hygromycin A; D017853:Phosphotransferases (Alcohol Group Acceptor); C063242:hygromycin-B kinase