Detection And Analysis Of Ac/Ds Transposon In Transgenic Soybean

One of the most effective ways in functional genomics research is the application of mutants. By comparing mutants with the control (CK), it is possible to know gene's function. Therefore, the construction of mutant pools and collection of various elite mutants are the basis for functional genomics research in soybean. Transposon tagging is a powerful molecular bio-technique for the construction of mutant pool and the study of gene function. This effective strategy has been applied in Arabidopsis successfully, and also has made great progress in rice in recent years. Whereas, there is little known about its application in soybean.The project was sponsored by the national high-tech program (973 Program). The aim of the thesis is to find out some homozygous lines by molecular biology methods. Such as PCR, Paromomycine Test, Southern Dot Blot, Kanamycin Test, RT-PCR, and DNA sequencing and so on. All the 940 T0 regeneration plants have already been detected, as well as the approximately 3000 T1 and T2 progenies. One possible mutant was found during the research and simple analysis was developed. Actually, this could be a foundation of gene cloning and soybean functional gene research. The following results are obtained:1,All the 940 T0 regeneration plants were detected by Paromomycine Test. It is supposed that there are some transgenic plants.2,PCR and Southern dot blot were carried out to detect the integration of nptâ…¡and Ac, Ds gene in T0 regeneration plants and the T1 and T2 progenies. And also, the transcription of Ac and Ds were detected by RT-PCR. There are seven possible Ac homozygous lines and nine possible Ds homozygous lines.3,Kanamycin can be used to select the transformed plants which have the nptâ…¡gene. In order to assay the optimal screening concentration, Kanamycin was smeared to the surface of the transformed soybean leaves of JilinXiaoli No.1.It showed that 500mg/L Km with 3 days selecting was most suitable.This method was compared with PCR as well, the results suggested that the equivalent ratio was 90.8% between the high-resistant plants and the resistant plants, 87.1%...