A convenient and efficient protocol for isolating high-quality RNA from latex of Hevea brasiliensis (para rubber tree)

Tang, Chaorong; Qi, Jiyan; Li, Heping; Zhang, Cunliang; Wang, Yuekun

doi:10.1016/j.jbbm.2007.04.002

Cited by 102 publications

(67 citation statements)

References 9 publications

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“…Sodium dodecyl sulfate (SDS) is used to harvest RNA from rubber-tree leaves and latex. 1,2) In our study, however, we found that RNA prepared by SDS-based methods from rubber-tree leaves, although suitable for common RT-PCR and Northern blot, was unsuitable for RACE in terms of integrity, since RACE performance makes greater demands on the integrity of RNA than common RT-PCR does. Other techniques, including cetyltrimethylammonium bromide (CTAB) [3][4][5][6] and TRIzol, 7,8) did not work well for rubber-tree leaves either.…”

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confidence: 63%

Extraction of High-Quality RNA from Rubber Tree Leaves

Deng

Luo

Zhang

et al. 2012

Bioscience, Biotechnology, and Biochemistry

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confidence: 63%

Extraction of High-Quality RNA from Rubber Tree Leaves

Deng

Luo

Zhang

et al. 2012

Bioscience, Biotechnology, and Biochemistry

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“…Cloning, expression and purification of HbCLP1, HbCLP2 and their domains Total RNA was isolated from the latex and young leaves of rubber trees (H. brasiliensis) as previously described [63]. Briefly, cDNA was synthesized, and the coding sequences of the chi-L1 (GenBank accession number KF648872) and chi-L2 (GenBank accession number KF648873) genes were amplified by PCR using the oligonucleotides CHI-a and CHI-b (all oligonucleotide sequences are detailed in Table S1).…”

Section: Methodsmentioning

confidence: 99%

Comparative study of two GH19 chitinase‐like proteins from Hevea brasiliensis, one exhibiting a novel carbohydrate‐binding domain

et al. 2014

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Plants express chitinase and chitinase-like proteins (CLPs) belonging to the glycosyl hydrolases of the GH18 and GH19 families, which exhibit varied functions. CLPs in the GH18 family have been structurally and functionally characterized; however, there are no structures available for any member of the GH19 family. In this study, two CLPs of the GH19 family from the rubber tree Hevea brasiliensis (HbCLP1 and HbCLP2) were cloned, expressed and characterized. HbCLP1 was identical to the allergen Hev b 11.0101 previously described by others, while HbCLP2 was a novel isoform exhibiting an unusual half chitin-binding domain before the catalytic domain. Sequence alignments showed that in the two proteins the catalytic residues Glu117 and Glu147 in HbCLP1 and HbCLP2, respectively, were mutated to Ala, accounting for the lack of activity. Nonetheless, both CLPs bound chitin and chitotriose (GlcNAc) 3 with high affinities, as evaluated with chitin-affinity chromatography and tryptophan fluorescence experiments. The chitin-binding domains also bound chitotriose with even higher affinities. The crystal structures of the HbCLP1-isolated domains were determined at high resolution. The analysis of the crystallographic models and docking experiments using (GlcNAc) 6 oligosaccharides provides evidence of the residues involved in sugar binding. Endochitinase activity was restored in both proteins by mutating residues A117E (HbCLP1) and A147E (HbCLP2); the distance between the catalytic proton donor and the catalytic nucleophile in the in silico mutated residues was 9.5 A, as occurs in inverting enzymes. HbCLP1 and HbCLP2 were highly thermostable and exhibited antifungal activity against Alternaria alternata, suggesting their participation in plant defense mechanisms. DatabaseThe atomic coordinates and structure factors have been deposited in the Protein Data Bank with accession numbers 4MST for the catalytic domain (CatD1) and 4MPI for the chitinbinding domain (CBD1). Nucleotide sequence data are available in the GenBank database under the accession numbers KF648872 (chi-L1) and KF648873 (chi-L2). AbbreviationsCatD1, catalytic domain of HbCLP1; CatD2, catalytic domain of HbCLP2; CatD, catalytic domain; CBD1, chitin-binding domain of HbCLP1; CBD2, chitin-binding domain of HbCLP2; CBD, chitin-binding domain; CLP, chitinase-like protein; GH, glycoside hydrolase; GlcNAc, Nacetylglucosamine; HbCLP, chitinase-like protein from Hevea brasiliensis; IPTG, isopropyl 1-thio-b-D-galactopyranoside; ITC, isothermal titration calorimetry; MES, 2-(N-morpholino)ethanesulfonic acid; NMR, nuclear magnetic resonance; PRP, pathogenesis-related proteins.

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“…Total RNA was extracted according to methods described in Tang et al, 2007. The quality and concentration of the extracted RNA were checked by agarose gel electrophoresis and measured by spectrophotometry (DU-70, Beckman, USA).…”

Section: Isolation Of Rnamentioning

confidence: 99%

Cloning and molecular characterization of a cDNA encoding a small GTPase from Hevea brasiliensis

Guo²,

Tian³

et al. 2013

Genet. Mol. Res.

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ABSTRACT. Small GTPases play a critical role in the regulation of a range of cellular processes including growth, differentiation, and intracellular transportation. The cDNA encoding a small GTPase, designated as HbGTPase1, was isolated from Hevea brasiliensis. HbGTPase1 was 882 bp long containing a 612-bp open reading frame encoding a putative protein of 203 amino acids, flanked by an 83-bp 5'-untranslated region (UTR) and a 187-bp 3'-UTR. The predicted molecular mass of HbGTPase1 is 22.62 kDa, with an isoelectric point of 5.06. The HbGTPase1 protein was predicted to possess the conserved functional regions of the small GTPase superfamily of proteins. Quantitative polymerase chain reaction analysis revealed that HbGTPase1 was constitutively expressed in all tissues tested. HbGTPase1 transcripts accumulated at relatively low levels in the flower, latex, and leaves, while HbGTPase1 transcripts accumulated at relatively high levels in bark. Transcription of HbGTPase1 in the latex was induced by jasmonate.

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A convenient and efficient protocol for isolating high-quality RNA from latex of Hevea brasiliensis (para rubber tree)

Cited by 102 publications

References 9 publications

Extraction of High-Quality RNA from Rubber Tree Leaves

Extraction of High-Quality RNA from Rubber Tree Leaves

Comparative study of two GH19 chitinase‐like proteins from Hevea brasiliensis, one exhibiting a novel carbohydrate‐binding domain

Cloning and molecular characterization of a cDNA encoding a small GTPase from Hevea brasiliensis

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