Molecular characterization and expression of buffalo (Bubalus bubalis) DEAD-box family VASA gene and mRNA transcript variants isolated from testis tissue

Ramakant Kaushik, Karn Pratap Singh, Vivek Bahuguna, K. Rameshbabu, Manoj Kumar Singh, Radhey Shyam Manik, Prabhat Palta, Suresh Kumar Singla, Manmohan Singh Chauhan

Research output: Contribution to journalArticlepeer-review

Abstract

VASA is a member of the DEAD-box protein family that plays an indispensable role in mammalian spermatogenesis, particularly during meiosis. In the present study, we isolated, sequenced, and characterized VASA gene in buffalo testis. Here, we demonstrated that VASA mRNA is expressed as multiple isoforms and uses four alternative transcriptional start sites (TSSs) and four different polyadenylation sites. The TSSs identified by 5'-RNA ligase-mediated rapid amplification of cDNA ends (RLM-5'-RACE) were positioned at 48, 53, 85, and 88 nucleotides upstream relative to the translation initiation codon. 3'-RACE experiment revealed the presence of tandem polyadenylation signals, which lead to the expression of at least four different 3'-untranslated regions (209, 233, 239 and 605 nucleotides). The full-length coding region of VASA was 2190. bp, which encodes a 729 amino acid (aa) protein containing nine consensus regions of the DEAD box protein family. VASA variants are highly expressed in testis of adult buffalo. We found five variants, one full length VASA (729 aa) and four splice variants VASA 2, 4, 5, 6 (683, 685, 679, 703 aa). The expression level of VASA 1 was significantly higher than rest of all (P. <. 0.05) except VASA 6. The relative ratio for VASA 1:2:4:5:6 was 100:1.0:1.6:0.9:48.

Original languageEnglish (US)
Pages (from-to)17-26
Number of pages10
JournalGene
Volume572
Issue number1
DOIs
StatePublished - Nov 1 2015
Externally publishedYes

Bibliographical note

Funding Information:
This work was funded by the National Agriculture Innovation Project (NAIP) grant to M.S.C. ( C-2067 and 075 ), ICAR, New Delhi, India. Author's thanks to Ms Neha Saini for providing language help.

Publisher Copyright:
© 2015 Elsevier B.V..

Copyright:
Copyright 2016 Elsevier B.V., All rights reserved.

Keywords

  • Isoform
  • RACE
  • Spermatogenesis
  • VASA

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