New candidate genes for sperm motility abnormality discovered by CGH microarray

Abstract

OBJECTIVE: The aim of this study is to investigate the genomic mutations occurred in the SR2d/SR2d male mice (whose spermatozoa are motionless and are thus infertile) using array comparative genomic hybridization (a-CGH) as an approach. DESIGN: Genomic mutations in the SR2d/SR2d mice are screened by comparing with its wild-type littermate mice by a-CGH. MATERIALS AND METHODS: Genomic DNA was isolated from testes of SR2d/SR2d and its wild-type littermate mice, labeled by cy-3 and cy-5 and hybridized to the pre-designed CGH microarray. Sperm motility-related high definition CGH microarrays (a total of 175,106 probes) have been designed based on the most updated Agilent database of about 4 million computationally validated CGH probes in this study. Some of the probes focus on the region from 30.3 Mb-39.7 Mb of chromosome 17 which is the region containing Mus Spretus chromatin in SR2/SR2 mice, the strain from which the SR2d/SR2d chromosome was derived. RESULTS: Three regions appear to be homozygously deleted by CGH analysis. These regions cover chromosome 17 from 38923583-39058450 bp, 39292540-39312921 bp and 40716449-40732825 bp respectively. All these three deletions are downstream of the SR2d/SR2d breaking site and are gene-poor. No known genes or RefSeq genes are located there. There are only 5 predicted genes (by Genscan, Geneid and Esembl, termed as 1174, 95281, 1178, 717 and 1192 respectively) discovered spanning the three deleted regions. PCR and RT-PCR are applied to confirm the genomic structure and the testicular expression of the 5 candidate genes respectively in the mutant mice and their wild-type counterparts. It has been discovered that all these five genes are expressed in the testis of wild-type mice whereas four of them (1174, 95281, 1178 and 717) are deleted in the sterile SR2d/SR2d male mice. CONCLUSIONS: The 4 deleted genes located on chromosome 17 discovered in this study could be important for the “motionlessness” in the spermatozoa.