Molecular Cloning And Expression Analysis Of A Novel Human Testis-specfic Gene, TDRG1

It is becoming increasingly clear that multiple genetic alterations or defects are responsible for male primary infertility most of which due to obstruction of spermatogenesis. Spermatogenesis is a complex process of cell development and differentiation. This process requires the highly regulated expression of multiple genes located on both the sex chromosomes and the autosomes. Discovery of spermatogenesis-related genes is a major challenge facing male infertility research. Cloning of new testis-specific gene related to spermatogenesis is of momentous physiological and pathological significance to understand the molecular mechanism of spermatogenesis.In this study, a new strategy Digital Differential Display (DDD) of the National Center for Biotechnology Information (NCBI), which is a statistical method to quantitate the transcript levels, was used to determine the fold differences between the libraries being compared. We have succeeded cloning a novel human testis-specific gene and studied the relationship of new gene and spermatogenesis with DDD combined with laboratory experiments such as RT-PCR,Northern blots,et al. There are two parts in this study, and the following are its main experimental methods and results:Part â…  Identification and expression of a novel human testis-specificgene TDRG1 by Digital Differential Display 1 Cloning in silico of new geneThe Unigene database of NCBI has thousands of expressed sequences, both known and novel, in the form of expressed sequence tags (ESTs), derived from diverse normal, pathological or tumor cDNA libraries, offer an attractive starting point for gene discovery. The DDD, a data-mining tool, takes advantage of the UniGene database by comparing a great number of ESTs from different libraries, which were assigned to a particular UniGene cluster. DDD was performed between 9 testis-derived EST libraries (pool A, or named "tester") and 76 different tissue-derived EST libraries (pool B, or named "driver"). Fold differences were calculated by using the ratio of pool A: pool B. Statistically significant...