转Bt cry1Ah基因抗虫玉米的遗传稳定性分析及农艺性状观察
发布时间：2019-04-09 11:37
【摘要】：在前期工作中,将Bt cry1Ah基因转入玉米自交系综31中,选取其中一株对亚洲玉米螟有显著抗性的玉米植株,命名为HGK60。分别从DNA水平和转录翻译水平对HGK60事件的T4代和T5代植株进行遗传稳定性分析。通过PCR、Tail-PCR、Southern Blot分析表明外源基因整合到了玉米基因组中并可以稳定遗传给后代,且明确了外源基因是按单插入双拷贝的方式插入到玉米1号染色体上,分析得到了玉米插入序列的侧翼序列。通过RT-PCR、Western Blot、ELISA、试纸条分析表明外源基因可以在玉米的根、茎、叶等组织中正常高效表达。在室内和田间分别用亚洲玉米螟、棉铃虫和黏虫幼虫对HGK60玉米的杀虫效果进行检测。室内生物活性检测结果表明,取食HGK60玉米心叶的亚洲玉米螟幼虫在3天后死亡率达100%;HGK60玉米对棉铃虫幼虫有毒杀作用,玉米不同部位对棉铃虫幼虫的杀虫效果存在差异;与综31相比,取食HGK60玉米叶片一周后的黏虫幼虫体重增长明显受到抑制。田间生物活性检测结果表明,HGK60玉米对于亚洲玉米螟和棉铃虫有很强的杀虫活性,达到高抗级别,对于黏虫有一定的杀虫活性,为抗性级别。对HGK60与郑58自交系回交转育5代后的转基因玉米进行农艺性状分析。在无虫害情况下,与郑58相比,HGK60的株高、穗位高、穗粗、穗长等差异不显著;在人工模拟虫害爆发的的情况下,郑58的生长发育受到影响,千粒重和单株产量都显著低于HGK60。以上研究表明,HGK60转基因抗虫玉米在玉米的抗虫育种上有很好的应用前景。
[Abstract]:In previous work, Bt cry1Ah gene was introduced into maize inbred ensemble 31, and one of the maize plants with significant resistance to Asian corn borer, named HGK60., was selected. The genetic stability of T4 and T5 generations of HGK60 events was analyzed at DNA level and transcriptional translation level, respectively. PCR,Tail-PCR,Southern Blot analysis showed that the exogenous gene was integrated into the maize genome and could be stably inherited to the progeny, and it was clear that the exogenous gene was inserted into maize chromosome 1 in the form of single insertion and double copy. The flanking sequence of maize insertion sequence was analyzed. The results of RT-PCR,Western Blot,ELISA, analysis showed that exogenous genes could be expressed normally and efficiently in roots, stems, leaves and other tissues of maize. The insecticidal effects of Asian corn borer, Helicoverpa armigera and Myxworm larvae on HGK60 maize were tested in laboratory and field, respectively. The results of laboratory bioactivity test showed that the mortality of Asian corn borer larvae fed on the heart leaves of HGK60 corn was 100% HGK60 after 3 days, and the insecticidal effects of different parts of corn on the larvae of cotton bollworm were different. Compared with Heizang 31, the weight gain of myxozoa larvae after feeding HGK60 maize leaves for one week was significantly inhibited. Field bioassay showed that HGK60 maize had strong insecticidal activity against Asian corn borer and Helicoverpa armigera, which reached high resistance level, and had certain insecticidal activity to Myxworm, which was the resistance level of corn borer and Helicoverpa armigera. The agronomic characters of transgenic maize after 5 generations of backcross between HGK60 and Zheng 58 inbred line were analyzed. Compared with Zheng 58, there was no significant difference in plant height, ear position, ear diameter and ear length between HGK60 and Zheng 58. Under the condition of artificial pest outbreak, the growth and development of Zheng 58 were affected, and the 1000-grain weight and yield per plant were significantly lower than that of HGK60.. The results indicated that HGK60 transgenic insect-resistant maize had good application prospect in maize pest-resistant breeding.
【学位授予单位】：西南科技大学
【学位级别】：硕士
【学位授予年份】：2016
【分类号】：S513
，
本文编号：2455147
[Abstract]:In previous work, Bt cry1Ah gene was introduced into maize inbred ensemble 31, and one of the maize plants with significant resistance to Asian corn borer, named HGK60., was selected. The genetic stability of T4 and T5 generations of HGK60 events was analyzed at DNA level and transcriptional translation level, respectively. PCR,Tail-PCR,Southern Blot analysis showed that the exogenous gene was integrated into the maize genome and could be stably inherited to the progeny, and it was clear that the exogenous gene was inserted into maize chromosome 1 in the form of single insertion and double copy. The flanking sequence of maize insertion sequence was analyzed. The results of RT-PCR,Western Blot,ELISA, analysis showed that exogenous genes could be expressed normally and efficiently in roots, stems, leaves and other tissues of maize. The insecticidal effects of Asian corn borer, Helicoverpa armigera and Myxworm larvae on HGK60 maize were tested in laboratory and field, respectively. The results of laboratory bioactivity test showed that the mortality of Asian corn borer larvae fed on the heart leaves of HGK60 corn was 100% HGK60 after 3 days, and the insecticidal effects of different parts of corn on the larvae of cotton bollworm were different. Compared with Heizang 31, the weight gain of myxozoa larvae after feeding HGK60 maize leaves for one week was significantly inhibited. Field bioassay showed that HGK60 maize had strong insecticidal activity against Asian corn borer and Helicoverpa armigera, which reached high resistance level, and had certain insecticidal activity to Myxworm, which was the resistance level of corn borer and Helicoverpa armigera. The agronomic characters of transgenic maize after 5 generations of backcross between HGK60 and Zheng 58 inbred line were analyzed. Compared with Zheng 58, there was no significant difference in plant height, ear position, ear diameter and ear length between HGK60 and Zheng 58. Under the condition of artificial pest outbreak, the growth and development of Zheng 58 were affected, and the 1000-grain weight and yield per plant were significantly lower than that of HGK60.. The results indicated that HGK60 transgenic insect-resistant maize had good application prospect in maize pest-resistant breeding.
【学位授予单位】：西南科技大学
【学位级别】：硕士
【学位授予年份】：2016
【分类号】：S513
，
本文编号：2455147
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