23 research outputs found
Co-extraction of high-quality RNA and DNA from rubber tree (Hevea brasiliensis)
High-quality nucleic acids are the basic requirement for performing genomic research. A reliable and efficient method was developed for co-extracting high-quality DNA and RNA from rubber tree (Hevea brasiliensis) in this study. Polyethylene glycol (PEG) and cetyltrimethylammonium bromide (CTAB) extraction buffer with high concentrations of polyvinylpyrrolidone (PVP) and ÎČ-mercaptoethanol was used in this study. The results show that 3.2% polyethylene glycol 8000 is the optimal concentration for successful separation of DNA and RNA. Spectrophotometric determination (A260/A280 and A260/A230 ratios), agarose electrophoresis analysis and reverse transcription (RT-PCR) of isolated nucleic acids indicate that high-quality DNA and RNA were extracted by this method. The general applicability of this method was also evaluated, and the results show that it was suitable for a variety of plants.Key words: Hevea brasiliensis, polyethylene glycol (PEG), nucleic acid, co-extraction, higher plants
A rapid method for isolation of low-molecular-weight RNA from Arabidopsis using low salt concentration buffer
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<w:LsdException Locked="false" Priority="37" Name="Bibliography" /> <w:LsdException Locked="false" Priority="39" QFormat="true" Name="TOC Heading" /> </w:LatentStyles> </xml><![endif]--> <!--[if gte mso 10]> <mce:style><! /* Style Definitions */ table.MsoNormalTable {mso-style-name:"Table Normal"; mso-tstyle-rowband-size:0; mso-tstyle-colband-size:0; mso-style-noshow:yes; mso-style-priority:99; mso-style-qformat:yes; mso-style-parent:""; mso-padding-alt:0cm 5.4pt 0cm 5.4pt; mso-para-margin:0cm; mso-para-margin-bottom:.0001pt; mso-pagination:widow-orphan; font-size:10.5pt; mso-bidi-font-size:11.0pt; font-family:"Calibri","sans-serif"; mso-ascii-font-family:Calibri; mso-ascii-theme-font:minor-latin; mso-fareast-font-family:ćźäœ; mso-fareast-theme-font:minor-fareast; mso-hansi-font-family:Calibri; mso-hansi-theme-font:minor-latin; mso-bidi-font-family:"Times New Roman"; mso-bidi-theme-font:minor-bidi; mso-font-kerning:1.0pt;} --> <!--[endif]--><span lang="EN-US">We have developed a rapid extraction method using low salt concentration buffer for the isolation of low-molecular-weight RNA from <em>Arabidopsis</em> tissues. The method was quick and efficient, and the small scale extraction process took no more than 1 hour, while yield and RNA quality were comparable with those of previously reported. The LMW RNA isolated using this method was high quality, abundant in small RNA and free of high molecular weight RNA. This method can be used to extract low-molecular-weight RNA for the purpose of small RNA cloning and detection, and library construction.</span
A Rapid and Economical Method for Low Molecular Weight RNA Isolation from a Wide Variety of Plant Species
Cloning, Expression Analysis, and Functional Characterization of Candidate Oxalate Transporter Genes of HbOT1 and HbOT2 from Rubber Tree (Hevea brasiliensis)
Secretion of oxalic acid from roots is an important aluminum detoxification mechanism for many plants such as Hevea brasiliensis (rubber tree). However, the underlying molecular mechanism and oxalate transporter genes in plants have not yet been reported. In this study, the oxalate transporter candidate genes HbOT1 and HbOT2 from the rubber tree were cloned and preliminarily identified. It was found that HbOT1 had a full length of 1163 bp with CDS size of 792 bp, encoding 263 amino acids, and HbOT2 had a full length of 1647 bp with a CDS region length of 840 bp, encoding 279 amino acid residues. HbOT1 and HbOT2 were both stable hydrophobic proteins with transmembrane structure and SNARE_assoc domains, possibly belonging to the SNARE_assoc subfamily proteins of the SNARE superfamily. qRT-PCR assays revealed that HbOT1 and HbOT2 were constitutively expressed in different tissues, with HbOT1 highly expressed in roots, stems, barks, and latex, while HbOT2 was highly expressed in latex. In addition, the expressions of HbOT1 and HbOT2 were up-regulated in response to aluminum stress, and they were inducible by metals, such as copper and manganese. Heterologous expression of HbOT1 and HbOT2 in the yeast mutant AD12345678 enhanced the tolerance to oxalic acid and high concentration aluminum stress, which was closely correlated with the secretion of oxalic acid. This study is the first report on oxalate transporter genes in plants, which provides a theoretical reference for the study on the molecular mechanism of oxalic acid secretion to relieve aluminum toxicity and on aluminum-tolerance genetic engineering breeding
Recent Advances in Flexible CNCâBased Chiral Nematic Film Materials
International audienceAbstract Cellulose nanocrystal (CNC) is a renewable resource derived from lignocellulosic materials, known for its optical permeability, biocompatibility, and unique selfâassembly properties. Recent years have seen great progresses in cellulose nanocrystalâbased chiral photonic materials. However, due to its inherent brittleness, cellulose nanocrystal shows limitations in the fields of flexible materials, optical sensors and food freshness testing. In order to solve the above limitations, attempts have been made to improve the flexibility of cellulose nanocrystal materials without destroying their structural color. Despite these progresses, a systematic review on them is lacking. This review aims to fill this gap by providing an overview of the main strategies and the latest research findings on the flexibilization of cellulose nanocrystalâbased chiral nematic film materials (FCNM). Specifically, typical substances and methods used for their preparation are summarized. Moreover, different kinds of cellulose nanocrystalâbased composites are compared in terms of flexibility. Finally, potential applications and future challenges of flexible cellulose nanocrystalâbased chiral nematic materials are discussed, inspiring further research in this field
<i>HbCBF1</i> overexpression in <i>Arabidopsis</i>.
<p>(A) Southern blotting of <i>HbCBF1</i> in transgenic plants. (B) <i>HbCBF1</i> expression in OE-3, OE-6 and OE-13 lines. The gene expression was detected by Northern blotting analysis using the <i>HbCBF1</i> probe. (C) Thirty days old seedlings of <i>col-0</i>, OE-3, OE-6 and OE-13 lines showed decreased plant height.</p
Gel mobility shift assay of recombinant HbCBF1 protein indicated CRT/DRE binding activity.
<p><b>(A)</b> Probes sequences used in this analysis. Bolded nucleotides are CRT/DRE motif, while underlined nucleotides are the mutation sites. <b>(B)</b> HbCBF1 bound with CRT/DRE elements specifically <i>in vitro</i>. Four microgram total proteins of GST (lane 1 from the left) or GST::HbCBF1 (lane 2â9 from the left) were used for the binding with labeled hot COR15a probe. One hundred and twenty-five folds molar unlabelled cold competitor probes were added as indicated (labe 3â9 from the left) in each reactions. The retarded binding bands and free non-binding probe were indicated as "B" and "F" respectively.</p
<i>HbCBF1</i> overexpression conferred cold resistance and relatively slow dehydration in <i>Arabidopsis</i>.
<p>(A) The electrolyte leakage rates of <i>col-0</i>, OE-3, OE-6 and OE-13 lines under nonacclimated (warm) or 5-days coldâacclimated conditions (cold). Electrolyte leakage was measured using leaves from two-week-old seedlings frozen to -7°C at a refrigerating rate of 1°C per h from -1°C. The star symbol (*) indicates significant difference between each individual OE line and wild type plants (WT). (B) Representative seedlings of <i>col-0</i>, OE-3, OE-6 and OE-13 lines recovered from 18 hours frozen at -6°C. The survival rates of each line were listed. (C) The dehydration rates of the detached leaves from <i>col-0</i>, OE-3, OE-6 and OE-13 seedlings. The star (*) symbol indicates significant difference between WT and each individual OE line, while plus symbols (+) shows significant difference between WT and individual OE-6 and OE-13 line. Data in A and C were presented as mean ± standard error of mean of five biological replicates. Significance was determined by Studentâs <i>t</i> test at the probability levels of <i>P</i> < 0.05. Comparisons were made between the wild type plant and each individual overexpressing line. For B, totally thirty to fifty plants were tested for each line and about ten seedlings were grown in each pot.</p