c-MYC mRNA is present in human sperm cells.
Stage-specific expression of proto-oncogenes, including c-myc, has been demonstrated during spermatogenesis in testis. Some of these proto-oncogenes are expressed postmeiotically, especially in the round spermatid stage. Recently, we demonstrated the presence of c-myc protein in mature ejaculated sperm cells with a possible role in sperm cell function. Since the half-life of c-myc protein has been shown to be short, we suspected the presence of c-myc mRNA in human sperm cells. In the present study, the presence of the c-myc mRNA transcript in human sperm cells was investigated by reverse transcriptase-polymerase chain reaction (RT-PCR) analysis and in situ hybridization. Total RNA, 5-10 micrograms, was extracted from 0.2-0.5 ml of pelleted human sperm cells by NP-40 lysis procedure, and was used to construct cDNA with pd(N)6 random primer and Moloney Murine Leukemia Virus (MMLV) reverse transcriptase. The PCR with sperm cDNA and primers #P1 and #P2, both from exon 3, resulted in amplification of the expected 322 bp product. Primers #P3 and #P4, which are located in exon 2 and exon 3, respectively, and are 1.37 kb apart, gave the expected PCR amplified 313 bp product ruling out the possibility of DNA contamination. The presence of c-myc mRNA in human sperm cells was further confirmed by in situ hybridization using a digoxigenin labelled DNA probe, containing exon 2 of the c-myc gene sequence. The c-myc specific DNA probe reacted with the postacrosomal mid-piece and tail regions of both noncapacitated as well as capacitated methanol-fixed sperm cells.(ABSTRACT TRUNCATED AT 250 WORDS)
Kumar
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Department of Molecular Biology and Genetics, Wayne State University, Detroit, MI 48201.
Patel
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Naz
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RK
D001483:Base Sequence; D015139:Blotting, Southern; D015342:DNA Probes; D015870:Gene Expression; D016259:Genes, myc; D006801:Humans; D017403:In Situ Hybridization; D008297:Male; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000096:biosynthesis*; D013091:Spermatogenesis; D013094:Spermatozoa / Q000378:metabolism*
D015342:DNA Probes; D012333:RNA, Messenger