The Preliminary Research On Transfering GMCBL Gene Into Soybean

Soybean (Glycine max (L.) Merr) is one of the most important plant protein sources for human diets and livestock feedings and contains essential amino acids for human dietary needs. The protein content of soybean exceeds that of the most commonly consumed food sources. Although soybean is an excellent source of protein, its nutritional quality is compromised by a low concentration of the sulfur amino acids methionine and cysteine. One of the potent ways to improve soybean sulfur-containing amino acid content is to manipulate the genes encoding critical enzymes involved in sulfate assimilation pathway by genetic engineering. Cystanthionine β-Lyase consents to be the second critical enzyme in the path of methionine biosynthesis, and also be the critical enzymein in the cysteine metabolic pathway. CbL plays an important role in metabolic pathway of sulfur-containing amino acid. Our results are listed as follows:1. In the former study of our lab, it has been proved that the transgenic tobacco of overexpressing GmCBL, which is located on chromosome03can improve the containing of Met. To get the higher expression soybean plants and improve soybean sulfur-containing amino acid content, we reconstructed the overexpressing vector of GmCBL containing a phosphinothricin resistance selectable marker, and induced GmCBL into soybean with by Agrobacterium-cotyledonary node transformation system.The result showed, no selective agent in SIM1,3-6mg/L glufosinate in SIM Ⅱ,and0.5-2mg/L glufosinate in SEM were the better choice. The selection pressure was determined by the different genetype.We got transplant102Nannong88-1,35Jack,11HZM. Through the identification of PCR, there are16nannong88-1,2Jack,1HZM.2. Some soybean varieties were transformed by pollen tube pathway, and their offspring were selected by the agents. In this research, the suitable concentrations of selection pressure are as follows:400mg/L Kana+0.10%Tween-20,150mg/L Basta+0.10%Tween-20,150mg/L Hyg+0.10%Tween-20in the first true leaf stage, and250mg/L Hyg+0.10%Tween-20in the compound leaf stage.In T2transgenic soybean plants, we got232plants of hyg-resistant from the vector of pMDC83-GmCBL,45plants of kana-resistant from the vector of pBI121-35S-GmOASTL4-gfp,45plants of kana-resistant from the vector of BI121-ap-GmOASTL4-gfp,28plants of Basta-resistant from the vector of pCambial300-BAR-GmDof4,62plants of Basta-resistant from the vector of pCambial300-BAR-GmDof11.