Establishment Of Site-specific Integrated And Marker-free Spider Silk-like Peptide Gene Transgenic Sheep Cell Line

Transgenic animal is the important material of modern biological research. However. the obtained transgenic animals have genetically modified selection of antibiotics and fluorescent marker genes which are the dangers to biological and environmental safety. Traditional genes randomly inserting into the genetically modified technology may cause abnormal expression of receptor species important gene, or lead to the expression silencing of exogenous gene.The study was to mediate gene fixed-point integration by phiC31 integrase, eliminate potential safety problems by the method of Cre/Loxp recombination enzyme system delete selection marker gene, to lay the establishment of cultivating no screening marker gene transfer to Spider Silk-like Peptide Gene 2S cloned sheep. Main results summarized as follows:1. PEGFP-N1 was used as skeleton carrier, containing attB sequence and two synthetic Loxp sequence of integration were successfully constructed in order to delete selection markers of eukaryotic expression vector pEGFP-N1-2S.2. phiC31 integrase mRNA and loaded with pEGFP-N1-2S expression of transfection sheep fetal skin fibroblasts in order to establish Spider Silk-like Peptide Gene adjustable point integration sheep cell line.3. The successful induction of Cre wear peptide membranes will be designated to remove selection marker gene of integrate sheep cell line, andthe fixed-point integration was established screening markers turn to silk protein gene 2S sheep cell line.