Agrobacterium-mediated transformation of durum wheat (Triticum turgidum L. var. durum cv Stewart) with improved efficiency

被引:71
作者
He, Y. [2 ]
Jones, H. D. [1 ]
Chen, S. [2 ]
Chen, X. M. [2 ]
Wang, D. W. [3 ]
Li, K. X. [1 ]
Wang, D. S. [2 ]
Xia, L. Q. [2 ]
机构
[1] Ctr Crop Genet Improvement, Harpenden AL5 2JQ, Herts, England
[2] Chinese Acad Agr Sci, Inst Crop Sci, Natl Key Facil Crop Gene Resources & Genet Improv, Beijing 100081, Peoples R China
[3] Chinese Acad Sci, Inst Genet & Dev Biol, State Key Lab Plant Cell & Chromosome Engn, Beijing 100101, Peoples R China
基金
英国生物技术与生命科学研究理事会;
关键词
Acetosyringone; Agrobacterium tumefaciens; durum wheat cv Stewart; picloram; transformation; FERTILE TRANSGENIC WHEAT; T-DNA DELIVERY; MICROPROJECTILE BOMBARDMENT; GENETIC-TRANSFORMATION; STABLE TRANSFORMATION; SELECTABLE MARKER; RAPID PRODUCTION; MATURE EMBRYOS; AESTIVUM L; CELL-CYCLE;
D O I
10.1093/jxb/erq035
中图分类号
Q94 [植物学];
学科分类号
071001 ;
摘要
An efficient Agrobacterium-mediated durum wheat transformation system has been developed for the production of 121 independent transgenic lines. This improved system used Agrobacterium strain AGL1 containing the superbinary pGreen/pSoup vector system and durum wheat cv Stewart as the recipient plant. Acetosyringone at 400 mu M was added to both the inoculation and cultivation medium, and picloram at 10 mg l(-1) and 2 mg l(-1) was used in the cultivation and induction medium, respectively. Compared with 200 mu M in the inoculation and cultivation media, the increased acetosyringone concentration led to significantly higher GUS (beta-glucuronidase) transient expression and T-DNA delivery efficiency. However, no evident effects of acetosyringone concentration on regeneration frequency were observed. The higher acetosyringone concentration led to an improvement in average final transformation efficiency from 4.7% to 6.3%. Furthermore, the concentration of picloram in the co-cultivation medium had significant effects on callus induction and regeneration. Compared with 2 mg l(-1) picloram in the co-cultivation medium, increasing the concentration to 10 mg l(-1) picloram resulted in improved final transformation frequency from 2.8% to 6.3%, with the highest frequency of 12.3% reached in one particular experiment, although statistical analysis showed that this difference in final transformation efficiency had a low level of significance. Stable integration of foreign genes, their expression, and inheritance were confirmed by Southern blot analyses, GUS assay, and genetic analysis. Analysis of T-1 progeny showed that, of the 31 transgenic lines randomly selected, nearly one-third had a segregation ratio of 3:1, while the remainder had ratios typical of two or three independently segregating loci.
引用
收藏
页码:1567 / 1581
页数:15
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