‘新陆早 33 号’棉花转基因体系建立及 AtPGIP1 基因的导入  

Establishment of Genetic Transformation System on Cotton(Gossypium hirsutum L.)Cultivar‘Xinluzao 33' and Transformation of AtPGIP1 Gene

作　　者：肖向文[1,2] 刘海峰 李雪源[3] 曹艳艳 王俊铎[3] 黄芳 郑巨云[3] 谢旗[4] 李晓波[1] 

机构地区：[1]中国科学院新疆理化技术研究所,干旱区植物资源化学重点实验室,乌鲁木齐830011 [2]中国彩棉(集团)股份有限公司,乌鲁木齐830016 [3]新疆农业科学院经济作物研究所,乌鲁木齐830091 [4]中国科学院遗传与发育生物学研究所,基因组生物学研究中心,北京100101

出　　处：《西北植物学报》2014年第4期658-664,共7页Acta Botanica Boreali-Occidentalia Sinica

基　　金：中国博士后科学基金(2012M521827);兵团博士资金(2013BB005);转基因特色专用棉新品种培育(2011ZX08005-005)

摘　　要：利用生物技术方法对棉花进行遗传改良主要限于有效的遗传转化系统。以新疆主栽优良陆地棉品种‘新陆早33号’为材料,利用下胚轴作为外植体对影响农杆菌介导的棉花遗传转化及体细胞胚胎发生的因素进行研究,成功建立了除草剂Basta筛选的棉花遗传转化技术体系。同时将植物抗病相关基因多聚半乳糖醛酸酶抑制蛋白基因AtPGIP1导入棉花,经过对再生转化植株的PCR鉴定,初步证明外源基因已经整合到棉花基因组。研究发现:Basta是棉花遗传转化中很有效的筛选剂,低浓度Basta(2.5mg/L)就能够获得很好的筛选效果;较低的共培养温度(20℃)及合适的农杆菌浓度(OD600=0.5)有助于提高转化效率。该研究结果表明,‘新陆早33号’具备作为棉花优良遗传转化受体的基本特征,研究中获得的15株AtPGIP1转基因植株经PCR分子检测均为阳性植株。该研究为新疆棉区棉花分子生物学研究及转基因育种研究奠定了重要基础。Genetic improvement in cotton via biotechnology is limited due to lack of an efficient transforma- tion and regeneration system. A reliable and high-efficiency system of genetic transformation mediated by Agrobacterium tumefaciens was developed in cotton （Gossypium hirsutum L. ） cultivar ＇Xinluzao 33＇ （an elite cultivar in Xinjiang area）, using hypocotyl as explants. Various aspects of transformation were exam- ined in efforts to improve the efficiency of producing transgenic plants. Meanwhile, a foreign gene Polygalacturonase-inhibiting protein （AtPGIP1）,which is related to plant disease resistance,was mauceu to mJ, cultivar. The integration of the interest gene and BAR transgenes into the genome of putative transgenic plants was confirmed by polymerase chain reaction （PCR）. Our results showed Basra is an effective selec- tion agent in cotton genetic transformation, which can be applied at a low concentration of 2. 5 mg/L. Agrobacterium at a relatively low concentration （OD600 =0.5） helped to improve the efficiency of transfor- mation. Relatively low co-cultivation temperature （20 ~C） was optimal for obtaining a higher efficient transformation. This result suggested that ＇Xinluzao 33 ~ can be used as a good receptor variety in cotton genetic transformation mediated by A. tumefaciens, increasing the range of cotton genotypes that can be transformed. Using this protocol, more than 15 normal plants were obtained through transformation of At- PGIPI,of which all regenerated plants were transgene-positive confirmed by PCR analysis. The protocol established in this study can be used in cotton functional genomics and in improving cotton cultivars by genetic engineering.

关 键 词：棉花 组织培养 体细胞胚胎发生 遗传转化 

分 类 号：Q789[生物学—分子生物学]