摘要
本研究建立了以甘蓝型油菜下胚轴为外植体的一步不定芽再生培养和遗传转化体系。首先,以甘蓝型油菜中双11号含部分子叶节的下胚轴为外植体,从6-BA和NAA配比、Ag NO3以及无菌苗苗龄等方面对影响油菜组织培养的因素进行了研究,建立了甘蓝型油菜快速高频一步不定芽再生培养技术体系。该技术体系为,切取5 d苗龄含部分子叶节的下胚轴置于添加4 mg/L 6-BA的MS基本培养基中培养,5 d再生出不定芽,再生频率为100%。在此基础上,进行了甘蓝型油菜的遗传转化,成功地将用p FGC5941双元载体构建的Bn TFL1基因干扰载体转入中双11号中。整个转化过程中,芽的诱导生成只需5 d,接着完成抗性芽的PPT筛选需要30 d,整个转化周期从播种到得到生根抗性苗仅需大约70 d,而传统转化方法抗性芽的诱导筛选需要90 d,整个转化周期需要130 d左右,大大缩短了转化周期,简化了转化过程,提高了转化效率。
In this study, we established a one-step shoot regeneration and transformation system using hypocotyls of Brassica napus as explants. We used hypocotyls including part of cotyledonary node from Zhongshuang 11 of Brassica napus L. as explants, and adjusted some elements influencing tissue culture of rapeseed such as hormone combinations of 6-BA and NAA, AgNO3 and seedling age, to establish one-step shoot regeneration system. The content of this system is that hypocotyls including part of cotyledonary node from five day seedlings were cultured in the MS media supplemented with 4 mg/L 6-BA, the shoots from the cultured explants were regenerated after five day culture later with the 100% frequency of shoot regeneration. Based on the above shoot regeneration system, a transformation experiment was carried out and a BnTFL1 RNAi vector constructed from pFGC5941 vector was transformed into Zhongshuang 11 successfully. In the whole transformation procedures, it takes only 5 days for inducing buds, and it spends 30 days to screen resistant buds by using PPT, which shorten 60 days of traditional transformation （90 days）. The whole transformation period from seeding to getting rooted resistant seedlings should span about 70 days that saves 60 days compared to traditional transformation period. Therefore, this new transformation method obviously shortened transformation period, simplified transformation process and improved transformation efficiency.
出处
《分子植物育种》
CAS
CSCD
北大核心
2015年第4期793-799,共7页
Molecular Plant Breeding
基金
湖南省科技计划重点项目(2011FJ4023)资助
关键词
油菜
下胚轴
子叶节
一步再生培养
遗传转化
Rapeseed
Hypocotyl
Cotyledonary node
One-step shoot regeneration
Transformation
