[Molecular cloning of a novel human testis-specific gene TDRG1].
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).


CONCLUSIONS
DDD combined with laboratory validation is an efficient method for identifying new human functional genes.
Yin
Guang-ming
GM
Department of Urology, Third Xiangya Hospital, Central South University, Changsha 410013, China. yingm75@126.com
Yang
Jian-fu
JF
Jiang
Xian-zhen
XZ
Tang
Yu-xin
YX
He
Le-ye
LY
Jiang
Zhi-qiang
ZQ
Zhong
Kuang-biao
KB
Zeng
Qing
Q
D000328:Adult; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D030541:Databases, Genetic; D005786:Gene Expression Regulation; D018398:Homeodomain Proteins / Q000235:genetics*; D006801:Humans; D008297:Male; D008969:Molecular Sequence Data; D009928:Organ Specificity; D011506:Proteins / Q000235:genetics*; D062085:RNA, Long Noncoding; D020133:Reverse Transcriptase Polymerase Chain Reaction; D016415:Sequence Alignment; D013737:Testis / Q000378:metabolism*
D018076:DNA, Complementary; D018398:Homeodomain Proteins; D011506:Proteins; D062085:RNA, Long Noncoding; C000603254:TDRG1 long non-coding RNA, human