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Obtaining Wheat ( Triticum aestivum L.) Lines with Yeast Genes for Trehalose Biosynthesis
Cytology and Genetics ( IF 0.5 ) Pub Date : 2020-08-06 , DOI: 10.3103/s0095452720040088
A. Yu. Kvasko , S. V. Isayenkov , K. V. Dmytruk , A. A. Sibirny , Ya. B. Blume , A. I. Yemets

Abstract

Yeast (Saccharomyces cerevisiae) genes for trehalose biosynthesis (TPS1 and TPS2) were transferred into genomes of several common wheat cultivars using two methods of Agrobacterium-mediated transformation (in vitro and in planta) to enhance drought tolerance. For this purpose, vectors pBract214-TPS1 and pBract214-TPS2 were constructed using the Gateway-cloning technique. Both the vectors contained TPS1 and TPS2 genes under control of the constitutive maize ubiquitin promoter (PUbi) and selectable marker hygromycin-phosphotransferase (hpt) gene. Three- to five-day-old calluses obtained from immature wheat embryos were used as explants for the transformation in vitro. Selection of transgenic plants was carried out on nutrient medium supplemented with 30 mg/L hygromycin (as a selectable agent). Seeds of wheat (transgenic generation T1) were obtained by the in planta method of transformation. Integration and the presence of yeast genes in wheat genomic DNA isolated from transgenic plants were confirmed by PCR analysis using primers specific to TPS1 and TPS2 genes.


中文翻译:

获得具有酵母基因的海藻糖生物合成小麦(Triticum aestivum L.)品系

摘要

使用两种农杆菌介导的转化方法(体外和植物),将用于海藻糖生物合成(TPS1TPS2)的酵母(酿酒酵母)基因转移到几个普通小麦品种的基因组中。为此目的,使用网关克隆技术构建了载体pBract214-TPS1和pBract214-TPS2。两种载体均包含在组成型玉米泛素启动子(PUbi)和选择标记潮霉素磷酸转移酶(hpt)控制下的TPS1TPS2基因。)基因。从未成熟小麦胚获得的三至五天大的老茧用作外植体用于体外转化。在补充了30 mg / L潮霉素(作为选择剂)的营养培养基上进行转基因植物的选择。通过植物内转化方法获得小麦种子(转基因T1代)。通过使用TPS1TPS2基因特异的引物进行PCR分析,确认了从转基因植物中分离的小麦基因组DNA中酵母基因的整合和存在。
更新日期:2020-08-06
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