[Detection of gonadotropin releasing hormone (GnRH) messenger RNA in the ovary and cloning of LH, chorionic gonadotropin (CG) receptor].
Detection of gonadotropin releasing hormone (GnRH) mRNA in rat ovary was carried out, and cloning of LH, chorionic gonadotropin (CG) receptor in human ovary was attempted by use of polymerase chain reaction (PCR). Firstly, in an attempt to detect the expression of GnRH or related gene in rat ovaries at the RNA level, GnRH message was amplified. Total RNA from rat ovaries was converted to cDNA using reverse transcriptase and amplified in PCR using a pair of specific primers complementary to the rat GnRH cDNA. The DNA products were subcloned into plasmid vectors and their sequence determined. 1. In the rat ovary, a prominent PCR product of 462 bp was identified as a fragment of prothymosin alpha cDNA previously found in the spleen. 2. In contrast, RT-PCR amplification of hypothalamus and granulosa cell messages indicated the presence of a 244 bp product with identical sequence to GnRH. To confirm the presence of GnRH message, a second set of GnRH primers was used. PCR amplification of cDNA from hypothalamus, granulosa cells and whole ovary yielded a product identical with the authentic GnRH cDNA sequence. These data demonstrated the presence of mRNA for GmRH and prothymosin alpha in the rat ovary. Secondly, a part of the human LH, CG receptor was obtained from human granulosa cells by selective amplification with PCR of DNA segments presenting possible sequence similarity with genes for the porcine or rat LH, CG receptor. Total RNA from human granulosa cells was converted to cDNA and amplified in PCR using degenerate oligonucleotide primers corresponding to possible conserved regions in extracellular segments of the porcine or rat LH, CG receptor.(ABSTRACT TRUNCATED AT 250 WORDS)
Oikawa
M
M
Department of Obstetrics and Gynecology, Hokkaido University School of Medicine, Sapporo, Japan.
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D003001:Cloning, Molecular; D004247:DNA / Q000235:genetics*; D005260:Female; D007987:Gonadotropin-Releasing Hormone / Q000032:analysis* / Q000235:genetics; D006801:Humans; D008969:Molecular Sequence Data; D010053:Ovary / Q000378:metabolism*; D016133:Polymerase Chain Reaction; D011498:Protein Precursors / Q000032:analysis / Q000235:genetics; D012333:RNA, Messenger / Q000032:analysis*; D051381:Rats; D011919:Rats, Inbred Strains; D011967:Receptors, Gonadotropin / Q000235:genetics*; D011974:Receptors, LH / Q000235:genetics*; D013947:Thymosin / Q000031:analogs & derivatives / Q000032:analysis / Q000235:genetics
D011498:Protein Precursors; D012333:RNA, Messenger; D011967:Receptors, Gonadotropin; D011974:Receptors, LH; C040782:prothymosin alpha; D007987:Gonadotropin-Releasing Hormone; D013947:Thymosin; D004247:DNA