Isolation and expression of an ischaemia-induced gene from gerbil cerebral cortex by subtractive hybridization.

Abstract

A subtraction cDNA library was made using subtractive hybridization of cDNA libraries constructed from gerbil cerebral cortex of control animals and animals 8 h after 10 min of transient forebrain ischaemia. After differential screening, a cDNA clone (named pGSH3) was isolated as a gene which is expressed only after the ischaemia insult. The cDNA insert of pGSH3 (0.7 Kb) hybridized to 2.8 Kb mRNA of ischaemic cerebral cortex. The gene was normally expressed in a small amount in the cerebellum, kidney, and lung, but was not expressed in the cerebral cortex, heart, liver, or jejunum in a detectable amount. Eight hours after the 10-min transient forebrain ischaemia, the gene expression became prominent in the cerebral cortex, and the amount of the mRNA also increased in the lung and kidney. An analysis of DNA sequence revealed that the pGSH3 insert has a 91.3 per cent homology with a 70-Kd human heat shock protein (HSP70) gene. These results indicate that an ischaemia-induced gene was isolated as a cDNA clone (pGSH3) by subtractive hybridization and differential screening. Expression of the gene was first detected in other organs especially in the kidney and lung after transient forebrain ischaemia.