基于PRLR-JAK-STAT5信号传导通路测定鹅催乳素的新方法

doi:10.11843/j.issn.0366-6964.2016.07.011

Abstract

Abstract:

In order to develop a high sensitive method of detecting goose prolactin，luciferase detecting system based on PRLR-JAK-STAT5 signal transduction pathway was designed in the present study.Firstly the CDS region of goose PRLR gene was cloned，the signal transducer and activator of transcription 5（STAT5) was artificially synthesized and inserted into the eukaryotic expression vector pCMV6-Entry and luciferase reporter pGL3-Enhancer plasmid as signal receiver vector and signal responder vector respectively.Secondly，the two recombinant vectors along with puromycin screening vector pEZX-MR03 and internal control pRL-TK vector were transfected into HEK293T cells.After puromycin screening，stable transgenic cell lines were isolated and stimulated by a serial of different concentrations of chicken PRL (0，30，60，90 ng•mL-1).Finally the mRNA relative expression level and enzyme activity of luciferase enzyme were detected by qRT-PCR and dual-luciferase detection system.Results showed that 10 stable transgene cell lines carried the 4 vectors were isolated.After treated with different concentrations of chicken PRL，luciferase gene mRNA expression level was up regulated and luciferase enzyme activity also enhanced in dose dependent manner in one of these 10 cell lines.These results demonstrated that it was feasible of detecting PRL bioactivity using the constructed PRLR-JAK-STAT5 signal transduction system and could be further used in measuring poultry PRL concentrations.

Key words: goose prolactin receptor, signal transduction, signal transducer and activator of transcription 5, luciferase reporter gene, prolactin bioactivity

CLC Number:

S835.2

SONG Jin-wei，LI Hui，CHEN Zhe，SHI Zhen-dan，WANG Zhen-yong. A Novel Method of Detecting Goose Prolactin Based on PRLR-JAK-STAT5 Signal Transduction Pathway[J]. ACTA VETERINARIA ET ZOOTECHNICA SINICA, 2016, 47(7): 1389-1395.

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A Novel Method of Detecting Goose Prolactin Based on PRLR-JAK-STAT5 Signal Transduction Pathway

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