title: Identification of differentially expressed genes of primary spermatocyte against round spermatid isolated from human testis using the laser capture microdissection technique.
abstract: The method of laser capture microdissection (LCM) combined with suppressive subtractive hybridization (SSH) was developed to isolate specific germ cells from human testis sections and to identify the genes expressed during differentiation and development. In the present study, over 10,000 primary spermatocytes and round spermatid cells were successfully isolated by LCM. Using the cDNAs from primary spermatocytes and round spermatids, SSH cDNAs library of primary spermatocyte-specific was constructed. The average insert size of the cDNA isolated from 75 randomly picked white clones was 500 bp, ranging from 250 bp to 1.7 kb. Using the dot-blot method, a total of 421 clones were examined, resulting in the identification of 390 positive clones emitting strong signals. Partial sequence of cDNAs prepared from each clone was determined with an overall success rate of 84.4%. Genes encoding cytochrome c oxidase II and the rescue factor-humanin were most frequently expressed in primary spermatocytes, suggesting their roles involved in meiosis.
authors.affiliation: National Laboratory Medical Molecular Biology, Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences, Peking Union Medical College, 100005 Beijing, China.
authors.forename: Gang
authors.initials: G
authors.lastname: Liang
authors.forename: Xiao Dong
authors.initials: XD
authors.lastname: Zhang
authors.forename: Lu Jing
authors.initials: LJ
authors.lastname: Wang
authors.forename: Yu Shen
authors.initials: YS
authors.lastname: Sha
authors.forename: Jian Chao
authors.initials: JC
authors.lastname: Zhang
authors.forename: Shi Ying
authors.initials: SY
authors.lastname: Miao
authors.forename: Shu Dong
authors.initials: SD
authors.lastname: Zong
authors.forename: Lin Fang
authors.initials: LF
authors.lastname: Wang
authors.forename: S S
authors.initials: SS
authors.lastname: Koide
mesh_terms: D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000032:analysis / Q000235:genetics* / Q000302:isolation & purification; D003576:Electron Transport Complex IV / Q000378:metabolism; D018507:Gene Expression Regulation, Developmental; D015723:Gene Library; D006652:Histological Techniques; D006801:Humans; D017403:In Situ Hybridization / Q000379:methods; D007834:Lasers; D008297:Male; D042282:Microdissection / Q000295:instrumentation / Q000379:methods*; D016133:Polymerase Chain Reaction; D012313:RNA / Q000235:genetics; D017422:Sequence Analysis, DNA / Q000379:methods; D013087:Spermatids / Q000737:chemistry* / Q000166:cytology / Q000378:metabolism; D013090:Spermatocytes / Q000737:chemistry* / Q000166:cytology / Q000378:metabolism; D013091:Spermatogenesis / Q000235:genetics; D013737:Testis / Q000166:cytology*
chemical_list: D018076:DNA, Complementary; D012313:RNA; C117108:cytochrome C oxidase subunit II; D003576:Electron Transport Complex IV