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

Authors

  • M. Z. S. AHMED Plant Genetic Resources Department, Desert Research Center (DRC) - 1, Mathaf El-Matarya Street, B.O.P 11753 El-Matarya, Cairo, Egypt
  • N. A. K. RASHED Plant Genetic Resources Department, Desert Research Center (DRC) - 1, Mathaf El-Matarya Street, B.O.P 11753 El-Matarya, Cairo, Egypt
  • A. AGORIO Institut des Sciences du Vegetal (ISV), Centre National de la Recherché Scientifique (CNRS), Paris, France
  • S. FILLEUR Institut des Sciences du Vegetal (ISV), Centre National de la Recherché Scientifique (CNRS), Paris, France
  • N. A. ABDALLAH Department of Genetics, Faculty of Agriculture, Cairo University, Giza, Egypt
  • A. A. HEMEIDA Genetic Engineering and Biotechnology Research Institute (GEBRI), Sadat City University (formerly Menoufia University), Egypt
  • M. I. NASR Genetic Engineering and Biotechnology Research Institute (GEBRI), Sadat City University (formerly Menoufia University), Egypt

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).

References

Ahmed, Z. F., A. M. Rizk, F. M. Hammouda and M. M. Seif El- Nasr (1972). Phytochemical Investigation of Egyptian Capparis species. Planta Med., 21: 156-160.

Apse, M. P., G. S. Aharon, W. A. Snedden and E. Blumwald, (1999). Salt tolerance conferred by overexpression of a vacuolar Na+/H+ antiport in Arabidopsis. Sci., 285: 1256-1258.

Ashoub, A., M. Knoblauch, W. Peters and A. Bel (2006). A simple extraction method for RNA isolation from plants. Egypt. J. Genet. Cytol., 35: 187-194.

Blumwald, E., G. S. Aharon and M. P. Apse (2000). Na+ transport in plant cell. Biochem. Biophys. Acta, 1465: 140-151.

Brunner, A. M., I. A. Yakovlev and S. H. Strauss (2004). Validating internal controls for quantitative plant gene expression studies. BioMed Central (BMC) Plant Biol., 4: 14-20.

Cherouana, S., A. Touil and S. Rhouati (2013). Two flavonoid glycosides from Lycium arabicum. Chem. Nat. Comp., 49: 930-931.

Fukuda, T., J. Yokoyama and H. Ohashi (2001). Phylogeny and biogeography of the genus Lycium (Solanaceae): Inferences from chloroplast DNA sequences. Mol. Phylogen. and Evol., 19: 246-258.

Gaxiola, R. A., R. Rao and A. Sherman (1999). The Arabidopsis thaliana proton transporters, AtNHX1 and Avp1, can function in cation detoxification in yeast. Proc. Nat. Acad. Sci. USA, 96: 1480-1485.

Guan, B., Y. Hu, Y. Zeng, Y. Wang and F. Zhang (2011). Molecular characterization and functional analysis of a vacuolar Na+/H+ antiporter gene (HcNHX1) from Halostachys caspica. Mol. Biol. Rep., 38: 1889- 1899.

Hasegawa, P. M., R.A. Bressan, J. K. Zhu and H. K. Bohnert (2000). Plant cellular and molecular rsponses to high salinity. Annu. Rev. Plant Physiol. Plant Mol. Biol., 51: 463- 499.

Hassanean, H., K. Desoky and A. El- Hamouly (1993). Quinovic acid glycosides from Zygophyllum album. Phytochem., 33: 663-666.

Hecker, K. H. and H. K. Roux (1996). High and low annealing temperature increase both specificity and yield in touchdown and stepdown PCR. BioTechniques, 20: 478-485.

Hirokawa, T., S. Boon-Chieng and S. Mitaku (1998). SOSUI: classification and secondary structure prediction system for membrane proteins. Bioinfor., 14: 378-379.

Kiefer, E., W. Heller and D. Ernst (2000). A Simple and efficient protocol for isolation of functional RNA from plant tissues rich in secondary metabolites. Plant Mol. Biol. Rep., 18: 33-39.

Korbie, J. D. and J. S. Mattick (2008). Touchdown PCR for increased specificity and sensitivity in PCR amplification. Nat. Protoc., 3: 1452-1456.

Li, J. Y., F. C. Zhang, J. Ma, L. Cai, Y. G. Bao and B. Wang (2003). Using RT-PCR to amplify the NHX gene fragment in Atriplex dimorphostegia. Plant Physiol. Commun., 39: 585-588.

Li, J., X. He, L. XU, J. Zhou, P. Wu, H. Shou and F. Zhang (2008). Molecular and functional comparisons of the vacuolar Na+/H+ exchangers originated from glycophytic and halophytic species. J. Zhejiang Uni. Sci., (B), 9: 132-140.

Liu, L., Y. Zeng, X. Pan and F. Zhang (2012). Isolation, molecular characterization and functional analysis of the vacuolar Na+/H+ antiporter genes from the halophyte Karelinia caspica. Mol. Biol. Rep., 39: 7193- 7202.

Ma, X. L., Q. Zhang, H. Z. Shi, J. K. Zhu, Y. X. Zhao, C. L. Ma and H. Zhang (2004). Molecular cloning and different expression of a vacuolar Na+/H+ antiporter gene in Suaeda salsa under salt stress. Biologia Plantarum, 48: 219 -225.

Moustafa, A. M. Y., A. I. Khodair, F. M. Hammouda and A. H. Husseiny (2007). Phytochemical and toxicological studies of Zygophllum album L. f.,. J. pharm. and Toxic., 2: 220-237.

Omar, S. A. S., Y. Al-Mutawa and S. Zaman (2007). Vegetation of Kuwait, Kuwait. Kuwait Inst. Sci. Res., 32-159.

Portillo, M., C. Fenoll and C. Escobar (2006). Evaluation of different RNA extraction methods for small quantities of plant tissue: Combined effects of reagent type and homogenization procedure on RNA quality, integrity and yield. Physiologia Plantarum, 128: 1-7.

Rauf, M., K. Shahzad, R. Ali, M. Ahmed, I. Habib, S. Mansoor, G. Berkowitz and N. Saeed (2014). Cloning and characterization of Na+/H+ antiporter (LfNHX1) gene from a halophyte grass Leptochloa fusca for drought and salt tolerance. Mol. Biol. Rep., 41: 1669-1682.

Saitou, N. and M. Nei (1987). The neighbor- joning method: a new method for reconstructing phylogenetic trees. Mol. Biol. Evol., 4: 406-425.

Shahina, A. G. (1994). Capparaceae. In: Boca Raton (ed). USA: CRC Press, Inc., 73. Shi, H. and K. J. Zhu (2002). Regulation of expression of the vacuolar Na+/H+ antiporter gene AtNHX by salt stress and abscisic acid. Plant Mol. Bio., 50: 543-550.

Shi, H., F. J. Quintero, J. M. Paro and J. K. Zhu (2002). The putative plasma membrane Na+/H+ antiporter SOS1 controls long-distance Na+/H+ antiporter in plants. Plant Cell, 14: 465-477.

Täckholm, V. (1974). Student’s Flora of Egypt, 2nd ed. Beirut: Cairo University Cooperative Printing Company, 162.

Tang, R., C. Li, K. Xu, Y. Du and T. Xia (2010). Isolation, functional char acterization, and expression pattern of a vacuolar Na+/H+ antiporter gene TrNHX1 from Trifolium repens (L.). Plant Mol. Biol. Rep., 28: 102-111.

Tattersall, A. R. E., A. Ergul, F. AlKayal, L. DeLuc, C. J. Cushman and R. G. Cramer, (2005). Comparison of methods for isolating high-quality RNA from leaves of grapevine. Am. J. Enol. Vitic., 56: 400-406.

Thompson, J. D., D. G. Higgins and T. J. Gibson (1994). ClustalW: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position- specific gap penalties and weight matrix choice. Nucleic Acids Res., 22: 4673-4680.

Wang, J., K. Zuo, W. Wu, J. Song, X. Sun, J. Lin, X. Li and K. Tang (2003). Molecular cloning and characterization of a new Na+/H+ antiporter gene from Brassica napus. DNA Seq., 14: 351-358.

Wu, C. X., X. H. Gao, X. Q. Kong, Y. X. Zhao and H. Zhang (2009). Molecular cloning and functional analysis of a Na+/H+ antiporter gene ThNHX1 from a halophytic plant Thellungiella halophila. Plant Mol. Biol. Rep., 27: 1-12.

Wu, G. Q., J. J. Xi, Q. Wang, A. K. Bao, Q. Ma, J. L. Zhang and S. M. Wang (2011). The ZxNHX gene encoding tonoplast Na+/H+ antiporter from the xerophyte Zygophyllum xanthoxylum plays important roles in response to salt and drought. J. Plant Physiol., 168: 758-767.

Zhang, H. X. and E. Blumwald (2001). Transgenic salt-tolerant tomato plants accumulate salt in foliage but not in fruit. Nat. Biotech., 19: 765-768.

Zorb, C., A. Noll, S. Karl, K. Leib, F. Yan and S. Schubert (2005). Molecular characterization of Na+/H+ antiporters (ZmNHX) of maize (Zea mays L.) and their expression under salt stress. J. Plant Physiol., 162: 55-66.

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2018-01-12

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