Discovery of germ cell-specific transcripts by expressed sequence tag database analysis

Abstract

Objective: To identify transcripts whose expression is restricted to germ cells.

Design: Expressed sequence tags (ESTs) from unfertilized egg libraries were utilized to perform in silico subtraction and identify germ cell-specific transcripts.

Setting: Baylor College of Medicine, Houston, Texas.

Animal(s): C57BL/6J/129SvEv hybrid.

Intervention(s): Tissue harvesting from mice.

Main outcome measure(s): Identification of germ cell-specific transcripts.

Result(s): We have used the Unigene collection of mouse cDNA libraries to identify ESTs derived from unfertilized egg libraries. A total of 3,499 ESTs were identified from Knowles Solter and Ko unfertilized egg cDNA libraries. In silico subtraction identified 258 ESTs, which were found in these unfertilized egg libraries, but not in adult mouse tissue cDNA libraries. We performed reverse transcription polymerase chain reaction (RT-PCR) on multiple adult tissues with 43 selected ESTs and found 5 of them where expression was absent in heart, lung, liver, brain, spleen, stomach, intestines, kidneys, and uterus, but restricted to ovaries and testes. Three ESTs were further analyzed, and they were exclusively localized to the oocytes by in situ hybridization.

Conclusion: We have shown that utilization of publicly available ESTs from murine EST libraries is a simple and rapid in silico approach to the identification of transcripts preferentially expressed in germ cells.