Cloning and differential expression of Na,K-ATPase in Penaeus vannamei

Abstract

To better understand the adaptive strategies of osmotic and ionic regulation in Penaeus vannamei, a full-length cDNA sequence encoding the Na,K-ATPase alpha subunit was isolated and characterized by rapid amplification of the cDNA ends (RACE). The nucleotide sequence was 3953 bp in length and included a 300 bp 5′ untranslated region (UTR), 536 bp 3′ UTR with an AATAA canonical polyadenylation signal sequence and poly (A) tail, and a 3117 bp open reading frame (ORF) encoding a 1038 amino acid protein. The calculated molecular weight of the protein was 115.4 kDa with an estimated pI of 5.29. This sequence was submitted to the National Center for Biotechnology Information (NCBI) GenBank under accession number KF765670. A phenetic analysis showed that the Na,K-ATPase α-subunit of P. vannamei shared a high degree of identity with other species. The relevant mRNA was detected in all of the tissues examined by fluorescent quantitative real-time polymerase chain reaction. The expression level was higher in the gills and hepatopancreas, and lower in the guts, muscles and epithelia. Increased expression of Na,K-ATPase mRNA levels was observed in all of the tissues examined in response to salinity exposure, and there was a significant difference in the tissues of the gills and hepatopancreas (P < 0.05); however, no statistical difference was observed in the epithelia, guts and muscles compared with the controls (P > 0.05). The findings demonstrated that the Na,K-ATPase α-subunit is highly conserved in crustaceans, and differential expression in different tissues of P. vannamei was observed, indicating that the gene might have multiple functions in the shrimp.

Key words:

• Differential expression
• K-ATPase
• molecular cloning
• Na
• Penaeus vannamei
• RACE

Disclosure statement

No potential conflict of interest was reported by the authors.

Funding

The research was supported by grants from the National High Technology 14 Research & Development of China (863) (grant numbers 2012AA10A404-4, 2012AA092104, 2013AA092901 and 2013AA092902); Strategic Priority Research of the Chinese Academy of Sciences (grant number XDA11020202); the National Natural Science Foundation of China (grant numbers 41276113, 41276114 and 41006069); science and technology cooperation projects of Sanya (grant number 2013YD74); the Sanya Station Database and the Information System of CERN, the Knowledge Innovation of the Chinese Academy of Sciences (grant number SQ201218); the Open Fund of Key Laboratory for Ecological Environment in Coastal Areas, State Oceanic Administration (grant number 201304); Guangdong Province (grant number 2012B061500012); and the Natural Science Foundation of Guangdong Province, China (grant number S2011040002133).

Supplementary material (Figure S1)

The supplementary material for this article is available via the Supplemental tab of the article's online page at http://dx.doi.org/10.1080/17451000.2015.1024131

Editorial responsibility: Eric Thompson