Cloning of expansin genes in ramie (Boehmeria nivea L.) based on universal fast walking.

Gene cloning is the first step to study the expression profiles and functions of a particular gene; considerable cloning methods have been developed. Expansin, thought to involve in the cell-wall modification events, was not cloned in ramie (Boehmeria nivea L.), which is one of the most important bast fiber crops with little conducted molecular research, especially on its fiber development. Studying the expansin gene family will uncover its possible relationship with ramie fiber development and other growth events. As a result, five expansin genes were cloned with full-length and their sequence information was investigated. Additionally, the phylogenetic analysis was conducted, which suggested that the cloned genes belong to the α-subfamily, and these genes expressed differently during ramie fiber developmental process. In this study, we aimed to apply a strategy for cloning novel full-length genes from genomic DNA of ramie, based on using degenerate primers, touchdown polymerase chain reaction and universal fast walking protocols. By cloning five full-length expansin genes, we believe the polymerase chain reaction-based gene cloning strategy could be applied to general gene studies in ramie and other crops.

[1]  Thomas D. Schmittgen,et al.  Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) Method. , 2001, Methods.

[2]  T Gojobori,et al.  Codon usage tabulated from the GenBank Genetic Sequence Data. , 1988, Nucleic acids research.

[3]  W. Rychlik,et al.  OLIGO 7 primer analysis software. , 2007, Methods in molecular biology.

[4]  Zhilong Tian.  Cloning and Expression of Cellulose Synthase Gene in Ramie [ Boehme- ria nivea (Linn.) Gaud.]: Cloning and Expression of Cellulose Synthase Gene in Ramie [ Boehme- ria nivea (Linn.) Gaud.] , 2008 .

[5]  Huang Cui-rong.  Effect of ramie's growth period on its structure and character , 2005 .

[6]  Zhixiang Zhang,et al.  Cloning, characterization, and expression analysis of acyl-acyl carrier protein (ACP)-thioesterase B from seeds of Chinese Spicehush (Lindera communis). , 2014, Gene.

[7]  Tian Zhi.  Cloning and Expression of Cellulose Synthase Gene in Ramie [Boehme-ria nivea (Linn.) Gaud.] , 2008 .

[8]  M. Hanafey,et al.  Insertion-deletion polymorphisms in 3′ regions of maize genes occur frequently and can be used as highly informative genetic markers , 2002, Plant Molecular Biology.

[9]  Synan F. AbuQamar,et al.  A mutation in the expansin-like A2 gene enhances resistance to necrotrophic fungi and hypersensitivity to abiotic stress in Arabidopsis thaliana. , 2013, Molecular plant pathology.

[10]  D. Cosgrove,et al.  © 2005 BioMed Central Ltd Summary , 2005 .

[11]  Eunkyoo Oh,et al.  A Triple Helix-Loop-Helix/Basic Helix-Loop-Helix Cascade Controls Cell Elongation Downstream of Multiple Hormonal and Environmental Signaling Pathways in Arabidopsis[C][W] , 2012, Plant Cell.

[12]  W. Gelbart,et al.  Universal Fast Walking for direct and versatile determination of flanking sequence. , 2002, Gene.

[13]  Jichen Xu,et al.  Transgenic Tobacco Plants Overexpressing a Grass PpEXP1 Gene Exhibit Enhanced Tolerance to Heat Stress , 2014, PloS one.

[14]  Huan Wang,et al.  Identification of a Bombyx mori gene encoding small heat shock protein BmHsp27.4 expressed in response to high-temperature stress. , 2014, Gene.

[15]  N. Zhang,et al.  Isolation of high quality RNA and construction of a suppression subtractive hybridization library from ramie (Boehmeria nivea L. Gaud.) , 2010, Molecular Biology Reports.

[16]  M. Frohman,et al.  Rapid amplification of cDNA ends (RACE). , 2011, Methods in molecular biology.

[17]  K. Roux,et al.  High and low annealing temperatures increase both specificity and yield in touchdown and stepdown PCR. , 1996, BioTechniques.

[18]  Y. Kapulnik,et al.  Microarray analysis and functional tests suggest the involvement of expansins in the early stages of symbiosis of the arbuscular mycorrhizal fungus Glomus intraradices on tomato (Solanum lycopersicum). , 2010, Molecular plant pathology.

[19]  Kang Liu,et al.  Protein expression changes during cotton fiber elongation in response to low temperature stress. , 2012, Journal of plant physiology.

[20]  Steven G. Schroeder,et al.  Single Nucleotide Polymorphisms and Insertion–Deletions for Genetic Markers and Anchoring the Maize Fingerprint Contig Physical Map , 2006 .

[21]  L. Zagorchev,et al.  The Role of Plant Cell Wall Proteins in Response to Salt Stress , 2014, TheScientificWorldJournal.

[22]  L. Lijun,et al.  Characterization by Suppression Subtractive Hybridization of Transcripts That Are Differentially Expressed in Leaves of Anthracnose-Resistant Ramie Cultivar , 2011, Plant Molecular Biology Reporter.

[23]  A genomic walking method for screening sequence length polymorphism , 2006 .

[24]  Weidong Wang,et al.  Isolation and characterization of a buffalograss (Buchloe dactyloides) dehydration responsive element binding transcription factor, BdDREB2. , 2014, Gene.

[25]  Synan F. AbuQamar.  Expansins: Cell Wall Remodeling Proteins with a Potential Function inPlant Defense , 2014 .

[26]  J. Mattick,et al.  'Touchdown' PCR to circumvent spurious priming during gene amplification. , 1991, Nucleic acids research.

[27]  Ping Chen,et al.  De novo assembly and characterization of transcriptome using Illumina paired-end sequencing and identification of CesA gene in ramie (Boehmeria nivea L. Gaud) , 2013, BMC Genomics.

[28]  Jie Chen,et al.  Transcriptome profiling using pyrosequencing shows genes associated with bast fiber development in ramie (Boehmeria nivea L.) , 2014, BMC Genomics.

[29]  Youn-sung Kim,et al.  Down-regulation of the IbEXP1 gene enhanced storage root development in sweetpotato , 2012, Journal of experimental botany.

[30]  Daniel J. Park,et al.  Universal Fast Walking Applied to cDNA , 2004, Preparative biochemistry & biotechnology.

[31]  Junping Gao,et al.  RhNAC2 and RhEXPA4 Are Involved in the Regulation of Dehydration Tolerance during the Expansion of Rose Petals1[C][W][OA] , 2012, Plant Physiology.

[32]  A. Hirsch,et al.  The expression of MaEXP1, a Melilotus alba expansin gene, is upregulated during the sweetclover-Sinorhizobium meliloti interaction. , 2004, Molecular plant-microbe interactions : MPMI.

[33]  Jie Liu,et al.  Functional analysis of BpDREB2 gene involved in salt and drought response from a woody plant Broussonetia papyrifera. , 2014, Gene.

[34]  M. Deyholos,et al.  Microarray analysis of flax (Linum usitatissimum L.) stems identifies transcripts enriched in fibre-bearing phloem tissues , 2007, Molecular Genetics and Genomics.

[35]  M. Nei,et al.  MEGA5: molecular evolutionary genetics analysis using maximum likelihood, evolutionary distance, and maximum parsimony methods. , 2011, Molecular biology and evolution.

[36]  A. Pavesi,et al.  Cloning of two glutamate dehydrogenase cDNAs from Asparagus officinalis: sequence analysis and evolutionary implications. , 2000, Genome.

[37]  S. Henikoff,et al.  Consensus-degenerate hybrid oligonucleotide primers for amplification of distantly related sequences. , 1998, Nucleic acids research.

[38]  K. Jacobsson,et al.  Cloning of ligand-binding domains of bacterial receptors by phage display. , 1995, BioTechniques.

[39]  F. De Leo,et al.  Genome walking in eukaryotes , 2011, The FEBS journal.

[40]  H. Goodman,et al.  Isolation of the Arabidopsis ABI3 gene by positional cloning. , 1992, The Plant cell.

[41]  T Gojobori,et al.  Codon usage tabulated from the GenBank genetic sequence data. , 1991, Nucleic acids research.

[42]  G. Crooks,et al.  WebLogo: a sequence logo generator. , 2004, Genome research.

[43]  Sun Zhi-min.  Correlation Between Fiber Fineness and Expression of Bast Fiber Development Related Gene FB27 in Ramie , 2009 .

[44]  S. McQueen-Mason,et al.  A role for expansins in dehydration and rehydration of the resurrection plant Craterostigma plantagineum , 2004, FEBS letters.

[45]  W. Gelbart,et al.  A modified universal fast walking method for single-tube transposon mapping , 2007, Nature Protocols.

[46]  Daniel J. Cosgrove,et al.  Loosening of plant cell walls by expansins , 2000, Nature.

[47]  M. Pezzotti,et al.  Overexpression of PhEXPA1 increases cell size, modifies cell wall polymer composition and affects the timing of axillary meristem development in Petunia hybrida. , 2011, The New phytologist.

[48]  Timothy Rose,et al.  CODEHOP (COnsensus-DEgenerate Hybrid Oligonucleotide Primer) PCR primer design , 2003, Nucleic Acids Res..