title: In vitro silencing of myostatin gene by shRNAs in chicken embryonic myoblast cells.
abstract: RNA interference represents one of the potential mechanisms of regulation of gene expression. Selective downregulation of myostatin (MSTN), a member of transforming growth factor-β (TGF-β) superfamily and a negative regulator of myogenesis, has been demonstrated to enhance skeletal muscle growth. In this study, we studied short hairpin RNA (shRNA)-induced myostatin gene silencing in chicken embryonic myoblast cells using seven different shRNA-expressing constructs by reverse transcription-quantitative real time PCR (RT-qPCR). Myostatin-silencing efficiency of all shRNA constructs were first evaluated in human embryonic kidney cell line 293T (HEK293T) cells, where we observed 30-75.6% reduction in myostatin expression, followed by chicken embryo myoblast cells that revealed up to 55% reduction in myostatin expression along with upregulation of MyoD by 4.65-folds. Consistent with the earlier observations, the transfection of cells with plasmids led to significant increase in interferon responsive genes OAS1 and IFN β (2-112-folds), independent of myostatin silencing in both HEK293T and chicken embryonic myoblast cells. Our study suggests that apart from shRNA sequences, cell type-specific factors may play a significant role in determining the knockdown efficiency of shRNAs.
authors.affiliation: Dept. of Animal Biotechnology, College of Veterinary Science & Animal Husbandry, Anand Agricultural University, Anand, Gujarat, 388001, India.
authors.forename: Ajai K
authors.initials: AK
authors.lastname: Tripathi
authors.forename: Mansi K
authors.initials: MK
authors.lastname: Aparnathi
authors.forename: Amrutlal K
authors.initials: AK
authors.lastname: Patel
authors.forename: Chaitanya G
authors.initials: CG
authors.lastname: Joshi
mesh_terms: D000818:Animals; D002642:Chick Embryo / Q000378:metabolism; D002645:Chickens / Q000235:genetics* / Q000378:metabolism; D057809:HEK293 Cells; D006801:Humans; D017570:MyoD Protein / Q000235:genetics / Q000378:metabolism; D032446:Myoblasts / Q000378:metabolism*; D055435:Myostatin / Q000235:genetics* / Q000378:metabolism; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000235:genetics* / Q000378:metabolism; D014162:Transfection
chemical_list: D017570:MyoD Protein; D055435:Myostatin; D034741:RNA, Small Interfering