doi:

DOI: 10.3724/SP.J.1259.2015.00354

Chinese Bulletin of Botany (植物学报) 2015/50:3 PP.354-362

Studies of the Transient Expression and Transformation of Cloned Thermostable α-Amylase Genes from Bacillus licheniformis in Tobacco and Arabidopsis


Abstract:
The full-length thermostable α-amylase gene from a Bacillus licheniformis strain was cloned in a prokaryotic expression vector and a plant expression vector with GFP as reporter gene. Thermostable α-amylase was overexpressed in Escherichia coli by IPTG induction for 6 h at 28℃, protein bands at 55 kDa were detected with SDS-PAGE, and activity of the enzyme was confirmed by amylolysis experiments. The plant expression vector was transformed into tobacco via Agrobacterium tumefaciens mediated transformation. GFP expression was observed both in cytoplasm and vacuoles of the lower epidermis of tobacco by fluorescence microscopy. Staining of the tobacco leaf by I2-KI revealed that the thermostable α-amylase expressed in tobacco leaf had enzyme activity. Finally, the gene was introduced into Arabidopsis by the floral dip method and homozygous transgenic plants expressing α-amylase gene were identified. The results of this research provide further information for overexpression of thermostable α-amylase in transgenic plants.

Key words:Bacillus licheniformis;thermostable α-amylase gene;tobacco transient expression;Arabidopsis

ReleaseDate:2015-07-13 09:55:52



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