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Published on: September 10, 2017
[Molecular cloning of a novel human testis-specific gene TDRG1]
Guang-ming Yin1, Jian-fu Yang, Xian-zhen Jiang
1Department of Urology, Third Xiangya Hospital, Central South University, Changsha 410013, China. yingm75@126.com
Objective:
To clone a novel human testis-specific gene TDRG1.
Methods:
A new contig of expression sequence tags (ESTs) Hs.180197 was identified from the testis libraries using digital differential display (DDD) to screen the novel human testis-specific gene. To validate the use of bioinformatics approaches in gene discovery, the ESTs Hs.180197, which was predicted to be testis specific, was chosen for further study. Reverse transcriptase-polymerase chain reaction (RT-PCR) was performed on different normal tissues to identify the expression of Hs.180197 in human testis. Using bioinformatics methods and IMAGE cloning of this contig, the full-length cDNA sequence of the noval human gene was cloned.
Results:
This novel gene was 1197 bp in length, located in chromosome 6p21.1-p21.2. The sequence of the open reading frame was 504-806 bp, as confirmed by RT-PCR and sequencing in human testis. The cDNA encodes a novel protein of 100 amino acids with a theoretical molecular weight of 10 000 and isoelectric point of 6.81. The sequence shares no significant homology with any known protein in the databases. Semi-quantitative RT-PCR analysis of multiple tissues further showed that the novel gene was expressed specifically in adult human testis. Considering a possible relation of this novel gene with the function of human testis, we named this new gene TDRG1 (testis development related gene 1, GenBank accession number: DQ168992).
Conclusion:
DDD combined with laboratory validation is an efficient method for identifying new human functional genes.
