RACE法扩增梅花鹿β-防御素-1(siBD-1)全长cDNA

doi:10.16431/j.cnki.1671-7236.2017.03.003

Abstract

Abstract:

In order to obtain the full-length cDNA of sika β-defensin-1 (siBD-1),5'-RACE and 3'-RACE primers were designed according to the partial sequences of siBD-1 that obtained from this study group, cDNA ends (RACE) technology were used to amplify siBD-1 cDNA from the total RNA of tongue mucosa tissue in sika, and amplified products were cloned into pMD18-T vector and subjected to PCR, restriction endonuclease digestion and sequencing. The results indicated that the length of siBD-1 cDNA 5'- and 3'- end were about 172 and 299 bp. The full-length siBD-1 cDNA was 418 bp (GenBank accession No. HM588696.1) and includes an 5'-untranslated region (UTR) of 89 bp, a open reading frame (ORF) of 192 bp, a stop codon of TAA, 3'-UTR of 118 bp and Ploy(A)16. The sequence homology showed that siBD-1 shared the greatest identity (90.6%) with buffalo enteric β-defensin, the higher identity with cattle (EBD, LAP, TAP, BNBD-4), goats (GBD-1, GBD-2), reindeer (reBD-1), sheep (sBD-1, sBD-2) and camel (caBD-1), that were 83.2%, 83.1%, 87.3%, 87.0%, 87.5%, 87.5%, 84.4%, 79.9%, 77.1% and 70.5%,respectively,and correspondingly low identity were 60.3% and 72.4% with horse (hoBD-1), pig(pBD-1), the lowest was 16.0% with human (hBD-2). The mature peptide was consisted of 38 amino acids, and contained 9 positively charged residues.

Key words: sika deer; 5'-RACE; 3'-RACE; β-defensin-1 (siBD-1)

CLC Number:

S865.4⁺2

TANG Juan, JIN Xin, ZHANG Man, HOU Yong-yue, LI Jun-yan, TIAN Qiao-zhen, LIU Jiao, WANG Yun-he, YANG Yin-feng. Amplification of the Full-length cDNA of Sika β-defensin-1(siBD-1) with RACE Method[J]. , 2017, 44(3): 635-643.

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Amplification of the Full-length cDNA of Sika β-defensin-1(siBD-1) with RACE Method

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