Cloning and expression analysis of a zinc-regulated transporters (ZRT), iron-regulated transporter (IRT)-like protein encoding gene in Dendrobium officinale
(2.Institute of Medicinal Plant Development, Chinese Academy of Medical Sciences-Peking Union Medical College, Beijing, China 100193)
【Abstract】The zinc-regulated transporters (ZRT), iron-regulated transporter (IRT)-like protein (ZIP) plays an important role in the growth and development of plant. In this study, a full length cDNA of ZIP encoding gene, designed as DoZIP1 (Gen Bank accession KJ946203), was identified from Dendrobium officinale using RT-PCR and RACE. Bioinformatics analysis showed that DoZIP1 consisted of a 1 056 bp open reading frame (ORF) encoded a 351-aa protein with a molecular weight of 37.57 kDa and an isoelectric point (pI) of 6.09. The deduced DoZIP1 protein contained the conserved ZIP domain, and its secondary structure was composed of 50.71% alpha helix, 11.11% extended strand, 36.18% random coil, and beta turn 1.99%. DoZIP1 protein exhibited a signal peptide and eight transmembrane domains, presumably locating in cell membrane. The amino acid sequence had high homology with ZIP proteins from Arabidopsis, alfalfa and rice. A phylogenetic tree analysis demonstrated that DoZIP1 was closely related to AtZIP10 and OsZIP3, and they were clustered into one clade. Real time quantitative PCR analysis demonstrated that the transcription level of DoZIP1 in D. officinale roots was the highest (4.19 fold higher than that of stems), followed by that of leaves (1.12 fold). Molecular characters of DoZIP1 will be useful for further functional determination of the gene involving in the growth and development of D. officinale.
【Keywords】 Dendrobium officinale; zinc-regulated transporters iron-regulated transporter-like protein (ZIP); gene clone; sequence analysis; real time quantitative PCR;
Coleman J E.Zinc enzymes[J].Curr Opin Chem Biol,1998,2 (2 ):222 .
Pu Q, Li S Z, Li P. Biotechnology Bulletin, 2012 (10): 15 (In Chinese).
Xiao J X, Qi X X, Zhang S L. Chinese Journal of Applied Ecology, 2010, 21(8): 1974 (In Chinese).
Guerinot M L.The ZIP family of metal transporters[J].BBABiomembranes,2000,1465(1 /2):190 .
Grotz N,Fox T,Connolly E L,et al.Identification of a family of zinc transporter genes from Arabidopsis that respond to zinc deficiency[J].Proc Natl Acad Sci USA,1998,95 :7220.
López-Millán A,Ellis D R,Grusak M A.Identification and characterization of several new members of the ZIP family of metal ion transporters in Medicago truncatula[J].Plant Mol Biol,2004,54 :583 .
Astudillo C,Fernandez A C,Blair M W,et al.The Phaseolus vulgaris ZIP gene family:identification,characterization,mapping,and gene expression[J].Front Plant Sci,2013,4 :286 .
Li S Z,Zhou X J,Huang Y Q,et al.Identification and characterization of the zinc-regulated transporters,iron-regulated transporter-like protein(ZIP)gene family in maize[J].BMC Plant Biol,2013,13 (114 ):3 .
Tiong J W,Mc Donald G K,Genc Y,et al.Hv ZIP7 mediates zinc accumulation in barley(Hordeum vulgare)at moderately high zinc supply[J].New Phytol,2014,201 (1 ):131 .
Chen W R,Feng Y,Chao Y E.Genomic analysis and expression pattern of Os ZIP1,Os ZIP3,and Os ZIP4 in two rice(Oryza sativa L.)genotypes with different zinc efficiency[J].Russ J Plant Physl+,2008,55 (33 ):400 .
Jiang M, Wang J, Wen G S, et al. China Journal of Chinese Materia Medica, 2013, 38(4): 504 (In Chinese).
Chen X M, Wang C S, Yang J S, et al. Chinese Pharmaceutical Journal, 2013, 48(19): 1634 (In Chinese).
Li C, Si J P, Gao Y H, et al. China Journal of Chinese Materia Medica, 2014, 39(2): 209 (In Chinese).
Zhang G, Zhang D W, Zhao M M, et al. Chinese Pharmaceutical Journal, 2013, 48(19): 1614 (In Chinese).
Li B, Tang K, Zhang G, et al. Chinese Pharmaceutical Journal, 2012, 47(22): 1790 (In Chinese).
Zhang G,Song C,Zhao M M,et al.Characterization of an A-type cyclin-dependent kinase gene from Dendrobium candidum[J].Biologia,2012,67 (2 ):360 .
Pfaffl M W.A new mathematical model for relative quantification in real-time RT-PCR[J].Nucleic Acids Res,2001,29 :e45.
Kozak M.An analysis of 50-noncoding sequences from 699 vertebrate messenger RNAs[J].Nucleic Acids Res,1987,15 (20 ):8125.
Frech C,Chen N.Genome-wide comparative gene family classification[J].PLo S ONE,2010,5 (10 ):e13409.