Genetic engineering structure comprising a coding sequence of human G-CSF with the regulatory regions of bovine β-lactoglobulin gene and reporter gene EGFP
On the basis of previously obtained pβLgGCSF plasmid containing the coding sequence of the gene human granulocyte-colony stimulating factor ( G-CSF ) under the control of regulatory elements of the bovine β-lactoglobulin gene and pGEMTcmvEGFP plasmid, containing the gene for EGFP (modified green fluorescent protein) under the promoter cmv (cytomegalovirus) and the bovine growth hormone polyadenylation site ( BGH рolyA site ), recombinant plasmid pβLgGCSFcmvEGFP was designed. From the pβLgGCSF plasmid on the Xba I site a βLgGCSF fragment was cut out, which after purification was cloned into a vector pBluescript II SK(-) hydrolyzed on the same site. From pGEMTcmvEGFP plasmid with BsmB I ( Esp3 I) restrictases the fragment cmv-EGFP-BGH PolyA site ( cmvEGFP ) was cut and purified, which was cloned in plasmid pβLgGCS on the site Not I. The βLgGCSFcmvEGFP construction was cut with Cla I restrictase and used to study the integration of transgene at different stages of ontogenesis of laboratory and farm animals.