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| Using Cre/lox Site-specific Recombination System to Construct the Restoring Line of Tobacco Engineered Male Sterile Line Induced by TA29-Barnase Gene |
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Abstract According to the principle of hybrid production, a new approach to restore the fertility of engineered male sterility induced by Barnase gene was constructed using the Cre/lox site-specific recombination system, which was distinct from the Barnase/Barstar restoring approach. In this study, the pBinBarloxTABn plant expression vector with a TA29-Barnase gene expression cassette flanked by two directly oriented lox sites and Bar gene expression cassette was transformed into Wisconsin 38 by Agrobacterium tumefaciens-mediated transformation. The male sterile plants appeared anther thinness, unfull and poor pollen produced. The pollen was unnormal, crimpy, and lost the germination ability, no normally expanded fruits and seeds formated were observed after self-pollinated. However,the normally fruits and seeds were obtained after pollinated using pollens from pBinCre transgenic plant. The hybrids were produced by pollinated BN1 and BN6 male sterile plants using pollen from pBinCre transgenic plant, total 23 F1 progenies from BN1 ×C1 hybrid, 22 F1 progenies from BN6 ×C1 hybrid were analyzed the TA29-Barnase gene deletion by PCR method.The results indicated the TA29-Barnase gene expression cassette had been excised efficiently from the genome of F1 progenies those having both Cre and Bar genes in them, the TA29-Barnase gene deletion rate reached 100% for both the BN1 ×C1 and BN6 ×C1 cross combinations. Those progenies that TA29-Barnase gene deleted could flower and fruit normally ,indicated the male sterility had been restored.
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Received: 06 May 2009
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