ISOLATION AND SEQUENCE ANALYSIS OF A NOVEL PARTIAL VACUOLAR Na+/H+ ANTIPORTER cDNA FROM Capparis orientalis, Lycium shawii AND Zygophyllum album

Abstract

Egyptian native (nondomesticated) plant species from different families such as Capparis orientalis, Lycium shawii and Zygophyllum album were collected from North Western coast of Marsa Matrouh Governorate, Egypt. Plant vacuolar Na+/H+ antiporter candidate gene from tolerant plant species; several are localized on the tonoplast, plays an important role in several plant species (Halophytes and xerophytes) under abiotic stress. Then, once the genes will be identified from tolerant plant species, the overall goal of this study is to identify partial the vacuolar antiporter NHX1 candidate gene. According to NHX1 family homologous sequence conservative region; one degenerate oligonucleotide primer pair was used to amplify core (partial middle) fragment of cDNAs vacuolar Na+/H+ antiporter with about size of 600 bp, approximately. Touchdown PCR program (TD-PCR) of cDNAs were success to increase specificity, sensitivity and yield to amplify core cDNA of vacuolar Na+/H+ antiporter gene. Sequence analysis provided us with a novel partial fragment length of cDNAs about 548 bp, 557 bp and 557 bp of a novel CoNHX1, LsNHX and ZaNHX was deposited in GenBank database with NCBI, GenBank accession no. KJ452345.1, KJ452346.1 and KJ452347.1 and amino acid sequences about 182 a.a, 185 a.a and 185 a.a with GenBank accession no. AHY19036.1, AHY19037.1 and AHY19038.1, respectively. BLASTN of sequences result and phylogenetic relationship analysis indicated that all were clustered into the vacuolar Na+/H+ antiporter group. The deduced amino acid sequences showed high identities with other plant vacuolar-type Na+/H+ antiporters. Taken together, these results suggest that CoNHX, LsNHX and ZaNHX are new members of the vacuolar Na+/H+ antiporter family. The ultimate goal of this study provided a basic foundation information about a Novel partial vacuolar Na+/H+ antiporter gene to develop 5` and 3` RACE technique (Rapid amplification cDNA Ends).