| A couple of experiments were conducted to improve the agronomic traits of the transgenic Bt rice, c.v. Kemingdao, which was resistant to stem borers and leaf folders, including development of a new rapid DNA extraction method and its application in PCR-based detection of transgenes, the methods and effectiveness of marker gene (gusA and hpt) assisted selection, agronomical traits improvement and breeding of new high-yielding Bt transgenic rice lines, and the analysis of yield, insect-resistance and Bt toxin content of newly bred lines. Results were summarized as the followings:A simple protocol was established for rice genomic DNA rapid extraction from etiolated seedlings. It could significantly save the time, reduce the cost of DNA extraction, and the DNA extracted was proven suitable for PCR-based rapid detection of transgenes in the improvement of Bt rice.The GUS histochemical assay was applicable to leaf segment, endosperm-cuts and germinating seeds in vivo for accurate and stable selection of plants with transgenes in the progeny of Kemingdao crossed to conventional varieties. Comparative analysis showed that endosperm-cuts staining was more cost-effective, more widely applicable, and therefore suitable for selection of transgenic plants and lines.Seeds of Kemingdao and its transgenic progenies could germinate normally when soaked in hygromycin B solution (50 mg · L-1), and with little effect on seedling emergency. No toxic symptom was observed on transgenic rice plants when sprayed with hygromycin B solution (75 mg·L-1), while brown spots were shown in non-transgenic plant leaves. Both seed soakingand foliar spray could be used for screening of transgenic rice, but the former is not suitable for recording of transgene segregation, and the later might be subject to weather effect.Comparing the endosperm-cuts GUS assay and hygromycin B spray for the selection of transgenic rice, the former seemed more labor-intensive but lower for reagent cost, while the later was on the opposite, and less error-prone in practice.The yield of transgenic Bt rice was improved through cross breeding using Kemingdao or its transgenic derivatives as transgene donor parent. In the process of breeding, if selection was solely based on insect-resistance of plants in paddy fields, without testing for the crylAb or marker genes, there was a high possibility of fault selection, resulting in many non-transgenic plants at the end and an overall low breeding efficiency.Breeding using the marker gene aided selection was complicated and time-consuming in the early generations, but the population became smaller in advanced generations. Therefore, the workload of marker-assisted selection method was not higher than that of conventional method on the whole, and could result in significant increase of opportunity for selecting insect resistant lines.Application of marker gene assisted selection, widening of the genetic distance of parents, or crossing with high-yielding new varieties, and increasing the population size of breeding materials at earlier generations, could all lead to more chance of selecting high yielding transgenic lines, through breaking the possible linkage of inferior agronomy performance and the transgene in Kemingdao.Anther culture and backcrossing could be used for rapid production of homozygous transgenic lines from progenies of transgenic rice crossed to conventional varieties, and thus was an efficient strategy for theimprovement of transgenic Bt rice.The yield trials and field resistance tests proved that the potential yield of the newly-bred lines was higher than that of Kemingdao (KMD1) while keeping the resistance as KMD1. One line bred from this experiment showed comparable yield with that of the leading commercial high yielding variety Xiushui63 in Zhejiang province.Correlation analysis showed there was no significant correlations among yield, major agronomic traits, and Bt toxin content for the new transgenic Bt lines, which implied that the improvement of yield in transgenic rice would not impair their resistance t...
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