Enrichment of haploid spermatids in mice by flow sorting.

Xiao-yu Xia; Xiao-yu Xia; Chen-zhi Guo; Chen XU

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Enrichment of haploid spermatids in mice by flow sorting.

Author: Xiao-Yu XIA ¹ ; Xiao-Yu XIA ² ; Chen-Zhi GUO ³ ; Chen XU ²
Author Information

1. Department of Histology and Embryology, Shanghai Jiaotong University School of Medicine, Shanghai 200025, China. zpxiaxy@shsmu.edu.cn
2. Department of Histology and Embryology, Shanghai Jiaotong University School of Medicine, Shanghai 200025, China.
3. Department of Biochemistry and Molecular and Cell Biology, Shanghai Jiaotong University School of Medicine, Shanghai 200025, China.
Publication Type:Journal Article
MeSH: Animals; Cell Separation; methods; Flow Cytometry; methods; Haploidy; Male; Mice; Mice, Inbred ICR; Spermatids; cytology
From: National Journal of Andrology 2014;20(2):106-110
CountryChina
Language:Chinese
Abstract:
OBJECTIVETo establish an effective method for haploid spermatid enrichment by Hoechst 33342 staining and flow sorting.
METHODSMouse testicular monoplast suspension was prepared by two-step enzyme digestion, and the cells were incubated in the medium containing Hoechst 33342 and Verapamil. Haploid spermatids were separated and enriched according to their DNA content by flow sorting. The gene expressions in the spermatids of several histone-modified enzymes, including the histone acetylases (HAT) and histone deacetylases (HDAC), were examined by RT-PCR and compared with that in the HAT-inhibitor curcumin-treated counterparts.
RESULTSWe successfully enriched the haploid spermatids with high purity and further purified the round and elongated spermatids. RT-PCR results indicated the specificity of the expression of the HAT gene in the spermatids, and that it was influenced by curcumin.
CONCLUSIONFlow sorting can efficiently improve the purity of haploid spermatid enrichment, which helps a lot to elucidate the mechanisms of spermiogenesis.