Sidorov Vladimir, Gilbertson Larry, Addae Prince, Duncan David
Monsanto Company, 700 Chesterfield Parkway North, St. Louis, MO 63017, USA.
Plant Cell Rep. 2006 Apr;25(4):320-8. doi: 10.1007/s00299-005-0058-5. Epub 2005 Oct 27.
Efficient production of seedling-derived Type I callus was demonstrated for several corn genotypes including commercial inbred lines. Seeds were germinated on MS-based medium containing 10 mg l(-1) picloram and 3 mg l(-1) 6-benzylaminopurine, which induced the development of axillary buds in the area of coleoptilar node. Nodal sections of 7-10-day old seedlings were isolated, split longitudinally, and placed on callus induction medium supplemented with 2.2 mg l(-1) picloram and 0.5 mg l(-1) 2,4-dichlorophenoxyacetic acid. For lines L4 and L9 the frequency of embryogenic callus induction was 38-42% based on calli per split nodal section. Frequency of callus induction from split nodal sections of seeds germinated on media without growth regulators was 0-3%. Seedling-derived callus of five genotypes was used for Agrobacterium-mediated transformation. Two constructs containing the green fluorescence protein gene and genes for either neomycin phosphotransferase II or glyphosate selection were used in transformation experiments. Transformation frequency varied from 2 to 11% and about 60% of the T(0) plants had 1-2 copies of transgenes.
已证明包括商业自交系在内的几种玉米基因型能够高效产生源自幼苗的I型愈伤组织。种子在含有10 mg l(-1) 毒莠定和3 mg l(-1) 6-苄基腺嘌呤的基于MS的培养基上萌发，该培养基诱导胚芽鞘节区域腋芽的发育。分离7-10日龄幼苗的节段，纵向切开，并置于补充有2.2 mg l(-1) 毒莠定和0.5 mg l(-1) 2,4-二氯苯氧乙酸的愈伤组织诱导培养基上。对于L4和L9品系，基于每个切开的节段产生的愈伤组织，胚性愈伤组织诱导频率为38-42%。在没有生长调节剂的培养基上萌发的种子切开节段的愈伤组织诱导频率为0-3%。五种基因型的源自幼苗的愈伤组织用于农杆菌介导的转化。在转化实验中使用了两种构建体，一种含有绿色荧光蛋白基因，另一种含有新霉素磷酸转移酶II基因或草甘膦选择基因。转化频率在2%至11%之间变化，约60%的T(0)植株有1-2个转基因拷贝。
