Construction Of Mark Free Salt-tolerance And Herbicide-resistant Vectors In Rice

Selectable marker genes (SMGs) are essential to plant genetic engineering and for the developmentof transgenic crops. However, the most used SMGs encode proteins resistant to antibiotics or herbicidesand use negative selection. Because SMGs are integrated into the plant genome, there are concernsabout widespread occurrence of transgenes in novel ecosystems. An unprecedented debate hasaccompanied the development and commercialization of transgenic crops. Therefore, there is a need forthe development of techniques for the efficient production of ‘clean’ marker-free transgenic plants.Production of marker-free transgenic crops eliminates risk of horizontal gene transfer and could mitigatevertical gene transfer. In this study, five salt-tolerance genes BADH, SKC1, AtNHX1, DREB2A andDREB2ACA constructs were engineered under the control of a root-specific promoter rolD respectively,and other five salt-tolerance vectors were constructed, containing BADH, SKC1, AtNHX1, DREB2A andDREB2ACA respectively, under the control of a green organs and root tips specific promoter CsVMV.All of these vectors were mark-free vectors. The double-resistance vectors were constructed withbar/2mg2-EPSPS and bar/GAT, under the control of a green organs and root tips specific promoterCsVMV and constitutive promoter CaMV35S. PCR, reconstriction enzyme cleavage and sequencinganalysis results showed that thirteen vectors containing the salt-tolerance genes and herbicide-resistantwere successfully constructed.