Screening And Cloning Up-regulated Expressed Genes Of Antithyroid Drug-induced Agranulocytosis In Graves Disease Patients

Objective To screen and clone full length genes which were specifically up-regulated expressed in Graves disease (GD) patients, who developed agranulocytosis induced by antithyroid drug.Methods (1) Adopting the lymphocytes separation medium separated the monocytes of GD patients, who developed agranulocytosis and who were normal by using antithyroid drug (ATD), joined with the EB virus to carry on cell culture.Using the technique of SSH to construct a subtractive library containing differentially expressed genes'fragments. (2) Cloning and primarily analysising those differentially expressed ESTs. (3) Using the Smart RACE technique to obtain the full length sequence of up-regulated expressed genes.Result The human antithyroid drug-induced agranulocytosis in GD subtractive library with high subtractive efficiency was successfully constructed and got 34 different EST . 21 cDNA fragments from SSH showed low homology with any known gene of human beings by using BLAST (http://www.ncbi.nhl.gov/Blast), it's indicated that these 21 cDNA fragments were from novel genes which specifically up-regulated expressed in antithyroid drug-induced agranulocytosis in GD. All these EST were to forward Genbank database, and may provide new targets for the research of pathogenesis of antithyroid drug-induced agranulocytosis in GD. The full-length gene sequence of differentially expressed genes would be attained by using RACE technique.Conclusion Through using SSH, we had successfully constructed the human antithyroid drug-induced agranulocytosis in GD subtractive library. Those 34 cDNA fragments which had 21 novel ESTs we got by screening the subtractive library might be promot invasion of antithyroid drug-induced agranulocytosis in GD.