ESTABLISHMENT AND ANALYSIS OF SOMATOSTATIN RECEPTORS (SSTR2 AND SSTR5) KNOCKOUT MOUSE

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We subcloned genomic DNA fragment of somatostatin receptor, SSTR2 and SSTR5, with PCR using several sets of oligoprimers. 3.8Kb Fragment of SSTR2 and 4.2Kb fragment of SSTR 5 were obtained. Both fragment consist with small portion of 5'franking sequence and receptor coding regions. Neo-cassette was inserted into the coding region and the fragments were subcloned into the pMCtk vector with HSV-tK.Those vectors were transfected to the ES cells, then, clones were selected by Neo-resistance first followed by positive selection with acyclovir. After this step, 35 clones were selected and PCR with primers outside the homologously recombined site revealed that 3 out of 35 clones were successfully recombinated with trasfected DNA Fragment (two clones for SSTR2 and one for SSTR5). These 3 clones were injected to mouse embryo. We got heterozygous F0 judging from the fur patches, however, failed to get homozygous knockout mice in F1 generations.