DISTRIBUTION, CLONING AND AMPLIFICATION SEQUENCE ANALYSIS OF GnRH GENE IN CULTURED GASTRIC PARIETAL CELLS OF RATS / 解剖学报

ABSTRACT

Objective To study the distribution and sequence analysis of gonadotropin releasing hormone (GnRH) gene in cultured gastric parietal cells of rats. Methods The distribution of GnRH molecule and GnRH mRNA were observed out through immunohistochemical ABC methods and in situ hybridization methods in cultured gastric parietal cells of rats. After isolation of the total RNA from the parietal cells, RT PCR was conducted to obtain GnRH cDNA. Then, the products of PCR was purified, digested by the restriction enzyme of Hind Ⅲ and EcoR Ⅰ, and DNA fragments interests were cloned into pUC19 vector. The products of PCR were analyzed by sequenceing with Sanger's method after identified by PCR and digestion of restriction enzyme. Results Gastric parietal cells showed GnRH immunoreactivity, positive material was located in cytoplasm with negative nuclei. GnRH mRNA hybridized signals were also detected in cytoplasm with negative nuclei. The specific amplified band of GnRH mRNA was detected through agarose gel electrophoresis and gene sequence is identical to the GnRH which has been reported in rat hypothalamus.Conclusion Our data suggest that GnRH could be produced by gastric parietal cells of rats and may modulate physiological function of gastric parietal cells of rats.\;[