柔嫩艾美耳球虫真核起始因子5和5A基因的克隆表达及转录动态分析

doi:10.11843/j.issn.0366-6964.2014.12.016

Abstract

Abstract:

In order to study the functions of eukaryotic initiation factors (eIFs) of Eimeria tenella，the genes encoding eIF5 (EteIF5) and eIF5A (EteIF5A) were cloned and detected for evaluating their transcriptional profiling during E.tenella different developmental stages.The primers were designed based on the predicted ORF sequences from E.tenella (Houghton) whole genome data （http://www.eupathdb.org），and the ORFs of EteIF5 and EteIF5A were cloned from Guangdong strain of E.tenella sporulated oocyst total RNA by RT-PCR.The ORF sequences of EteIF5 and EteIF5A were inserted into the prokaryotic expression vector pMAL-c2x，and expressed in Escherichia coli，respectively.Five total RNA samples were extracted from 5 developmental stages of E.tenella，i.e.，unsporulated oocysts，sporulating for 7 hours，sporulated oocysts，sporozoites，and 2^ndgeneration merozoites，and subsequently were transcripted into the first strand of cDNA.The E.tenella β-actin gene was chosen as a control，and the transcriptional levels of EteIF5 and EteIF5A were detected by Real-time RT-PCR method.The sequencing results showed that cloned ORF sequences of EteIF5 and EteIF5A are 1 248 bp and 486 bp in their length，encoding the peptides composed of 315 and 161 amino acids，respectively.The qRT-PCR results revealed that EteIF5 and EteIF5A showed different transcription levels through the E.tenella developmental stages，and both of EteIF5 and EteIF5A in sporozoites were all higher than other stages.The cloning and expression profiling of EteIF5 and EteIF5A genes of E.tenella was studied firstly as we known，and these results will provide an important basis for probing the functions of E.tenella eIFs and be helpful to provide more insights into development of novel anti-coccidials.

Key words: Eimeria tenella, eIFs, gene cloning, expression profile, qRT-PCR

CLC Number:

S855.9

GOU Ling-qiao，GONG Zhen-xing，YU San-ke，YIN Hao，MA Xue-ting，CAI Jian-ping，LIN Qing. Cloning and Expression Profiling of Eukaryotic Initiation Factor 5 and 5A Genes in Eimeria tenella[J]. ACTA VETERINARIA ET ZOOTECHNICA SINICA, 2014, 45(12): 2019-2027.

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Cloning and Expression Profiling of Eukaryotic Initiation Factor 5 and 5A Genes in Eimeria tenella

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