Hypocotyl-based Agrobacterium-mediated transformation of soybean (Glycine max) and application for RNA interference.
An efficient system of gene transformation is necessary for soybean [Glycine max (L.) Merrill] functional genomics and gene modification by using RNA interference (RNAi) technology. To establish such system, we improved the conditions of tissue culture and transformation for increasing the frequency of adventitious shoots and decreasing the browning and necrosis of hypocotyls. Adding N(6)-benzylaminopurine (BAP) and silver nitrate in culture medium enhanced the shoot formation on hypocotyls. BAP increased the frequency of the hypocotyls containing adventitious shoots, while silver nitrate increased the number of shoots on the hypocotyls. As a result, the number of adventitious shoots on hypocotyls cultured in medium containing both BAP and silver nitrate was 5-fold higher than the controls. Adding antioxidants in co-cultivation medium resulted in a significant decrease in occurrence of browning and necrosis of hypocotyls and increase in levels of beta-Glucuronidase (GUS) gene expression. Histochemical assays showed that the apical meristem of hypocotyls was the "target tissue" for Agrobacterium tumefaciens transformation of soybean. Gene silencing of functional gene by using RNAi technology was carried out under above conditions. A silencing construct containing an inverted-repeat fragment of the GmFAD2 gene was introduced into soybean by using the A. tumefaciens-mediated transformation. Several lines with high oleic acid were obtained, in which mean oleic acid content ranged from 71.5 to 81.9%. Our study demonstrates that this transgenic approach could be efficiently used to improve soybean quality and productivity through functional genomics.
Wang
Geliang
G
Key Laboratory of Photosynthesis and Environmental Molecular Physiology, Institute of Botany, Chinese Academy of Sciences, 100093 Beijing, China.
Xu
Yinong
Y
D016960:Agrobacterium tumefaciens / Q000235:genetics*; D000975:Antioxidants / Q000494:pharmacology; D001593:Benzyl Compounds / Q000494:pharmacology; D015139:Blotting, Southern; D044943:Fatty Acid Desaturases / Q000235:genetics / Q000378:metabolism; D005227:Fatty Acids / Q000378:metabolism; D005966:Glucuronidase / Q000378:metabolism; D018546:Hypocotyl / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism; D018519:Meristem / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D008957:Models, Genetic; D018520:Plant Shoots / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D030821:Plants, Genetically Modified; D016133:Polymerase Chain Reaction; D011687:Purines / Q000494:pharmacology; D034622:RNA Interference; D012835:Silver Nitrate / Q000494:pharmacology; D013025:Glycine max / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism; D014170:Transformation, Genetic
D000975:Antioxidants; D001593:Benzyl Compounds; D005227:Fatty Acids; D011687:Purines; D012835:Silver Nitrate; D044943:Fatty Acid Desaturases; C040448:delta-12 fatty acid desaturase; D005966:Glucuronidase; C480551:benzylaminopurine