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由来自水稻成熟胚的愈伤组织或由此而建立的悬浮细胞系作基因枪转化的靶材料 ,将质粒 p ILTAB2 2 7(含 hph基因和 gus基因 )导入籼稻 ,在 Basmati- 1、青油粘、新山粘 2 9、胜优 2号 ,明恢 63等品种获得了转基因植株。 T0 转化体 gus基因组织化学染色和 DNA分子杂交 ( Southern blot)证实 ,gus和 hph基因已整合到上述品种的基因组中。本实验仔细研究了hph基因在 Basmati- 1的 T1、T2 和 T3等各个世代的遗传及表达。对 T1家系中各个体核 DNA的分子检测表明 ,hph基因普遍呈 3∶ 1的分离 (转基因阳性∶阴性 ) ,表明 hph基因为单一位点的整合 ,其遗传符合单 (显性 )基因控制的孟德尔遗传规律。还发现一些株系表现 1∶ 1的不正常的分离 (阳性个体较预期的少 ) ,推测这是由于雌或雄配子之一方的 hph基因受到抑制而产生基因沉默的结果。在 T2 、T3世代获得了具 hph基因的纯合体。笔者还探讨了外源基因在核基因组中的遗传稳定性及基因的沉默与活化问题
Plasmid pILTAB227 (containing the hph gene and the gus gene) was introduced into indica rice by callus transformed from the mature embryo of rice or the suspension cell line thus established as the target material for the gene gun transformation. In Basmati-1, Sticky, sticky Johor 2 9, Shengyou 2, Minghui 63 and other varieties obtained transgenic plants. T0 transformants gus histochemical staining and Southern blot analysis confirmed that the gus and hph genes have been integrated into the genomes of these breeds. In this experiment, we carefully studied the inheritance and expression of hph gene in various generations of Basmati-1 such as T1, T2 and T3. Molecular detection of individual nuclear DNA in T1 pedigrees showed that the hph gene was generally 3: 1 segregated (transgene positive: negative), indicating that the hph gene was a single site integration that was genetically conformed to the single (dominant) gene control Mendelian law of inheritance. Some lines were also found to exhibit an abnormal 1: 1 separation (less than expected for the positive individuals), presumably as a result of gene silencing resulting from the inhibition of the hph gene on one of the female or male gametes. In T2, T3 homozygotes with the hph gene were obtained. The author also explored the genetic stability of exogenous genes in the nuclear genome and gene silencing and activation issues