基因枪法导入cyclAt和GUS基因获得转基因水稻

Introduction of cyclAt and GUS into Rice to Obtain Transgenic Plants by Particle Bombardment

以cyclAt基因为目的基因,HPT和GUS为选择和报告基因,应用PDS-1 000/He型基因枪协同转导,通过轰击来源于水稻成熟种子诱导产生的胚性愈伤组织,获得了转基因水稻植株.结果表明:通过X-Gluc染色分析,GUS基因在愈伤组织和再生植株叶片中的表达效率高.以转基因植株的基因组DNA为模板,用cyclAt基因的特异性引物对其进行PCR扩增,只检测到1 000 bp左右的片段,比其1 604 bp的完整序列小,因此推测cyc1At基因在转导入水稻后被剪接.此外,在协同转导过程中,cyc1At和GUS可能与受体基因组DNA随机整合,因此视GUS基因为报告基因有一定局限性;但两者同时被整合的频率远高于单一基因的整合.

Target gene cyclAt, choice gene HPT and report gene GUS were co-transformed into the embryonic calli originated from mature rice seeds via particle bombardment method, transgenic rice plants were obtained. The X-Gluc staining analysis showed that report gene GUS expresses with high efficiency in both callus and leaf. Meanwhile, with the template of genome DNA of the transgenic plant leaves and the specific primers to amplify cyc 1 At by PCR, only a segment of 1 000 bp nucleotides was obtained, smaller than the fuU sequence of 1 604bp, so we conjectured that cyc 1 At has changed the configuration or has been montaged after being transformed into the acceptor rice. In addition, the results also showed that there exist limitations by using GUS as report gene to evaluate cyc 1 At transformation, for these two genes might integrate randomly into genome DNA of the acceptor, although the transformation frequency with both of them integrated simultaneously was indeed much higher than with any single gene integration.