摘要
本试验以茄子下胚轴和子叶为外植体,通过对抗生素选择压筛选、不同抗生素抑菌效果比较、农杆菌侵染时间、预培养时间、共培养时间等条件优化,建立了适用于茄子的遗传转化体系,并利用农杆菌介导法将抗根结线虫Bt cry6A基因导入茄子中,以期获得抗线虫的茄子材料,探索新的抗线虫育种途径,为利用茄子转基因技术培育和改良茄子品质提供理论依据。结果表明,75mg·L-1的卡那霉素可以有效地抑制愈伤组织的产生;200mg·L-1特美汀能够完全抑制农杆菌的生长;最佳侵染外植体为子叶,在预培养4d,侵染10min,共培养1d时转化率最高;获得的56株Kan抗性植株中有15株具有GUS活性;经过PCR、RT-PCR验证,初步确认Bt cry6A基因已整合至茄子基因组中,并在转录水平上正常表达;根结线虫幼虫接种鉴定表明,转Bt cry6A基因茄子再生植株对南方根结线虫抗性明显提高。
The eggplant genetic transformation system were established,through optimization of antibiotic selection pressure,effects of different antibiotic inhibitory,agrobacterium infection time,pre-culture time,and coculture time,using the hypocotyl and cotyledon as explants and agrobacterium mediated method. And the Bt cry6 A gene was integrated into the eggplant by agrobacterium-mediated transformation,so as to obtain the nematode-resistant eggplant and explore new ways of nematode-resistance breeding, and lay the foundation for the cultivation and improvement of quality of eggplant by the use of transgenic technology. The result showed that： the kanamycin of 75 mg·L^- 1could effectively inhibit the generation of callus; the timentin of 200mg·L^- 1could completely inhibit the growth of agrobacteria; and the best infection explant was cotyledon,whose conversion rate was the highest one by pre-cultured with 4d,infected with 10 min and co-cultured with 1d. There were 15 plantlets which had GUS activity in 56 regenerated plants resistant to kanamycin,Confirmed by PCR and RT-PCR verification,Bt cry6 A gene had been integrated into the eggplant genome and expressed respectively normally at the transcriptional level. Root-knot nematode inoculation identification showed that the resistance of transgenic eggplant with Bt cry6 A was improved compared with the control.
出处
《核农学报》
CAS
CSCD
北大核心
2015年第4期685-691,共7页
Journal of Nuclear Agricultural Sciences
基金
国家科技支撑项目(2012BAD02B02
2013BAD01B04-6)
关键词
茄子
植株再生
遗传转化
BT
cry6A基因
根结线虫
eggplant
plant regeneration
genetic transformation
Bt cry6A gene
root knot nematode
