Cloning And Analysis Of Genes Involved In Germination Of Young Soybean Embryo

Abstrect: soybean is important source of vegetable protein and plant oil ,and is an important cash crop.So it has been attached greatly to genetic improvement. Immature embryo culture technology in plant genetic breeding as an assisted breeding technology, not only can be used to prevent embryo abortion (immature embryo rescue), but also can be used to shorten the breeding cycle as accelerating the stability of plant traits. At the same time, the tchenique can be used for transgenic breeding and somaclonal variation breeding. Compared with other staple crops, soybean immature embryo culture technology is still quite backward. Therefore, it is an important approach to soybean genetic improvement by cultivation of soybean embryo. Cloning and analysis of genes related to embryo germination in soybean is also fancinating for its genetic improvement.In this study, soybean"ribenqing"immature embryos was used as the materials, and devided into two groups. One group was incubated on 100 rpm shaking, 37℃, no light conditions for 48 hours, another at 25℃for 48 hours under normal (as control). Total RNA of the two groups were inversed-transcripted into cDNA and then amplicated using 78 pairs of different primers. Results showed that most of these bands with difference between amplicants are 500bp about. The differential bands in gel were extracted. The extracts were amplified again and then separated by 1% agarose gel and ligased to clone vector. Host bacteria (E. coli) were transformed by ligased products. White colonies were selected blue white screening. Plasmids were extracted by alkalinelysis method. The insert was identified by EcoRⅠand HindⅢrestriction enzyme digestion.9 positive fragments have been screened out by Reverse Northern hybridization. The difference fragment K3, K4, K5, K7, K17, K19, K27, K28, K30 were sequenced by Shanghai Bioengineering Technology Service Co., Ltd.. The sequences were analysed by BLAST software. Results showed that K3 is homologous to the alpha subunit of elongation factor in protein biosynthesis homologous,and no homologous genes for other fragments have been found. The results sugested they have some function in the process of soybean young embryo germination.