不同甜瓜品种遗传转化再生体系的建立与基因编辑植株的快速获取  

Establishment of genetic transformation and regeneration systems for different melon varieties and rapid acquisition of gene edited plants

作　　者：卡地尔阿依·买买提 周婷婷 韩盛[1] 梅丽克汗·热西提 玉山江·麦麦提[1] Kadierayi Maimaiti;ZHOU Tingting;HAN Sheng;Meilikehan Rexiti;Yushanjiang Maimaiti(Key Laboratory of Integrated Pest Management on Crops in Northwestern Oasis,Ministry of Agriculture and Rural Affairs,P.R.China/Institute of Plant Protection,Xinjiang Academy of Agricultural Sciences,Urumqi 830091,China;Hami Agricultural Machinery Technology Promotion Service Center,Hami Xinjiang 839000,China)

机构地区：[1]新疆农业科学院植物保护研究所/农业农村部西北荒漠绿洲作物有害生物综合治理重点实验室,乌鲁木齐830091 [2]哈密市农业农机技术推广服务中心,新疆哈密839000

出　　处：《新疆农业科学》2024年第7期1666-1672,共7页Xinjiang Agricultural Sciences

基　　金：国家自然科学基金地区科学基金项目“基于CRISPR/Cas9技术的哈密瓜抗多病毒病非转基因材料创制研究”(31760511)。

摘　　要：【目的】研究不同甜瓜品种遗传转化再生体系的建立与基因编辑植株的快速获取。【方法】选用巴吾顿、老汉瓜、其里甘、新密杂11号(86-1)4个甜瓜品种的子叶为外植体,通过农杆菌介导法侵染甜瓜子叶,采用组织培养法建立老汉瓜的遗传转化再生体系,运用PCR法检测获取基因编辑植株。【结果】从4个甜瓜品种中筛选出老汉瓜为优势品种作为外植体,在MS+6-BA 1.0 mg/L+NAA 0.05 mg/L培养基预培养2 d和共培养3 d,在MS+6-BA 1.0 mg/L+NAA 0.05 mg/L+潮霉素5 mg/L+头孢霉素250 mg/L筛选培养基上继续培养7 d,在MS+6-BA 1.0 mg/L+NAA 0.05 mg/L+潮霉素5 mg/L+头孢霉素250 mg/L的不定芽诱导培养基上培养1~2周可见长出的小芽,将诱导芽分别放置于不加潮霉素和加潮霉素的芽伸长培养基上培养,诱导芽在不加潮霉素的芽伸长培养基上的生长速度比加潮霉素的培养基生长快,可使芽生长至2叶苗期缩短3周,在无菌环境下剪去一个芽提取DNA,并采用RT-PCR方法检测获得21株转化植株。【结论】筛选出基因编辑植株嫩芽,不加潮霉素的芽伸长培养基可将获得基因编辑植株周期缩短3周。【Objective】Study on establishment of genetic transformation and regeneration systems for different melon varieties and rapid acquisition of gene edited plants.【Methods】The cotyledons of four melon varieties,including Bawudun,Laohangua,Qiligan,and Xinmiza No.11(86-1).The sweet melon cotyledons were infected by Agrobacterium tumefaciens mediated method,and a genetic transformation and regeneration system of Laohan melon was established by tissue culture method and then genetically edited plants were obtained through PCR detection.【Results】The results showed that Laohan melon was selected as the dominant variety from four melon varieties as the explants.It was pre-cultured on MS+6-BA 1.0 mg/L+NAA 0.05 mg/L medium for 2 days and co-cultured for 3 days,then it was further cultured on MS+6-BA 1.0 mg/L+NAA 0.05 mg/L+hygromycin 5 mg/L+cephalosporin 250 mg/L screened medium for 7 days,after that,small buds were seen growing on MS+6-BA 1.0 mg/L+NAA 0.05 mg/L+hygromycin 5 mg/L+cephalosporin 250 mg/L adventitious bud induction medium for 1-2 weeks.The induced buds were placed on both non hygromycin and hygromycin added bud elongation medium for cultivation.It was found that the growth rate of the induced buds on the non hygromycin added bud elongation medium was faster than that on the hygromycin added medium,which shortened the growth of the buds to the two leaf seedling stage by 3 weeks.One bud DNA was cut off in a sterile environment and 21 transformed plants were examined by RT-PCR.【Conclusion】Gene edited plant buds were successfully selected on a shoot elongation medium with added hygromycin.The use of a shoot elongation medium without added hygromycin can shorten the time for obtaining gene edited plants by 3 weeks.

关 键 词：甜瓜 农杆菌介导 遗传转化 基因编辑植株 

分 类 号：S652[农业科学—果树学]