mRNA差异显示分离棉花抗旱耐盐基因的相关cDNA片段

Isolation of cDNA Fragment Related to Cotton Resistance to Drought and Salt Genes by mRNA Differential Display

采用mRNA差异显示(DDRT-PCR)技术,对抗旱、耐盐棉花品种(品系)分别进行25%PEG6000和1.3%NaCl胁迫诱导,利用优化的蛋白酶K法分别提取胁迫与非胁迫棉花叶片中的总RNA,以此为模板,选用12个锚定引物和20个随机引物的240个组合进行RT-PCR,产物经6%变性聚丙烯酰胺凝胶电泳后银染,共获得88条差异条带,二次PCR扩增,选择出41条表达稳定的差异条带。

Using mRNA differential display（DDRT-PCR） , the cotton variety of resistance to drought and salt is treated separately by 25 %PEG6000 and 1.3% NaC1. The total RNA of the treated and non-treated cotton leaves are drawed separately by using the way of improved proteinase- K pathway. The 240 combinations which are established by 12 anchored primers and 20 arbitrary primers are conducted by RT-PCR, the products are applied by the method of silver-staining with 6 % PAGE. 88 different displayed fragments are obtained, and 41 of which are picked up by the second PCR.