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ARS2,ARS7,ARS8是从水稻珍汕97A不育系核DNA中克隆的一组在酵母细胞中能起始 DNA复制的片段.其中ARS8是单拷贝顺序,而ARS2,ARS7却是高度重复顺序.将带有ARS2,ARS7,ARS8的质粒用低压脉冲电泳法导入水稻部分酶解小细胞团中,经浅层培养,辅板,获得愈伤组织,Km筛选获得抗性愈伤组织.抽提抗性愈伤组织DNA,经South-ern点杂交证实ARS2,ARS7分别有3.2%,3.6%愈伤组织导入了外源基因,较对照高1倍和 1. 3倍.HindⅢ酶解抗性愈伤组织 DNA,琼脂糖凝胶电泳分离,转膜,Southern杂交,发现ARS2和 ARS7呈多位点整合.
ARS2, ARS7 and ARS8 are clones that can initiate DNA replication in yeast cells cloned from the nuclear DNA of Zhenshan 97A CMS line. Among them ARS8 is a single copy sequence, while ARS2 and ARS7 are highly repetitive sequences. The plasmids with ARS2, ARS7 and ARS8 were introduced into partially digested rice cells by low voltage pulse electrophoresis. After the supernatant was cultured in shallow layer, the callus was obtained and the callus was screened by Km to obtain the resistant callus. The DNA of resistant callus was extracted. The Southern blot analysis showed that the exogenous genes of ARS2 and ARS7 were 3.2% and 3.6%, respectively. 3 times. Hind Ⅲ enzymatic digestion of the callus DNA, agarose gel electrophoresis separation, transfection, Southern hybridization and found ARS2 and ARS7 was multi-site integration.