cloning and expression of a human interferon a2 gene in e. coli

the plasmid palca1sifn containing cdna that encodes the human interferon a-2b was obtained from the atcc(no. 531667). in this system the expression of the gene is under the control of an alca promoter. alca p is a specific promoter for expression of different genes in aspergillusfilamentous. in this plasmid the coding region of ifn?-2b is preceded by the coding region of a synthetic signal peptide. for direct expression of the ifna-2b gene under the control of a t7/lac promotor of pet24d(+) expression vector, three subcloning steps were carried out which resulted in the construction of 3 new plamids. pha1, pha2 and pha4 in which the ifna-2 gene is under the control of lacp, lacuv5p and t7 promoter, respectively. another plasrnid, pha3, was also constiucted and is a modified version of pet24d(+). provided there is a synthetic signal peptide preceding the coding region of ifna2 gene, the protein can be seen in either system, as shown by western blotting, albeit with a different level of expression. the best product can be seen in the pha4plasmid with a t7 pas judged by dot and western blotting