| Because rabbits possess characters of physiology and anatomy closer to human race than mice, they are much superior than mice as animal models of human diseases like hypertrophic cardiomyopathy, perturbed lipoprotein metabolism and atherosclerosis. In additional, rabbits have been widely applied in the studies of angiocarpy system, pulmonary diseases, eye diseases, metabolism diseases, toxicity tests of new drugs, Acquied Immune Deficiency Syndrome (AIDS), and cancer. The use of live bioreactors for the expression of human genes in the mammary gland of transgenic animals is one of the most cost-effective ways for the production of valuable recombinant therapeutic proteins. Rabbit is a good kind of species for the expression of tens to hundreds of grams of therapeutic proteins in the milk during lactation. The lactating mammary gland of rabbits has proven to be effective in the processing of complex proteins. So rabbits are also good candidates for bioreactors.Recently our laboratory has successfully established the technology for somatic cells cloned rabbit, which makes it possible to produce rabbits with modifications to specific genes by the combination of homologous recombination and subsequent prescreening of nuclear donor cells. Our laboratory is building a technology platform for producing transgenic cloned rabbits.This study is dedicated to some basic work to somooth the way of establishing the transgenic cloned rabbits platform. Our work includes two parts. In the first part, we cloned rabbit HPRT gene using the recombineering system. We constructed a knockout HPRT targeting vector by replacing the last three exons with a PGK-Neo-bGHpA cassette; we also constructed a promoterless knockin targeting vector by inserting an IRES-EGFP-IRES-Neo-bGHpA cassette in the 3'UTR of HPRT gene. In the second part, we cloned rabbit POU5F1 gene and constructed a promoterless knockin targeting vectors by inserting an IRES-EGFP-IRES-Neo-bGHpA cassette in the 3'UTR of POU5F1 gene.
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