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EASYspin Plant RNA Kit

Manufactured by Aidlab
Sourced in China

The EASYspin Plant RNA Kit is a tool designed for the rapid and efficient extraction of high-quality RNA from plant tissues. It utilizes a specialized spin column-based approach to isolate RNA, allowing for a streamlined and reliable purification process.

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33 protocols using EASYspin Plant RNA Kit

Firstly, total RNA was extracted using the EASY spin Plant RNA Kit (Aidlab, Beijing, China). Then, HiScript® III-RT SuperMix for qPCR (Vazyme, Beijing, China) was used to synthesize the first-strand cDNAs. Snapgene software was used to design primers (Table S6), and the ABI-7500 Connect Real-Time PCR Detection System was used to perform qRT-PCR. With 1 μL of template in a reaction volume of 20 μL, cDNAs were diluted to 200 ng, and three technical repetitions were performed. The PCR amplification procedure was as follows: 95 °C for 30 s, 95 °C for 10 s, 60 °C for 30 s, and 60 °C for 15 s. GAPDH was used as the internal reference gene. Primer sequences are presented in Table S6.
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Total RNA from mixed tissues of the plant seedlings was extracted using an EASYSpin Plant RNA kit (Aidlab, Beijing, China), and the first-strand cDNA was synthesized by being reverse transcribed with a HiScript II 1st strand cDNA synthesis kit (Vazyme Biotech, Nanjing, China). The RT-PCR amplification enzyme (TB Green Premix Ex TapII), primer and template were mixed and amplified in a CFX96-RT-PCR machine. Three biological replicates were generated for each sample, and the results were calculated by the 2−ΔΔCT method. The primers used for qRT-PCR are listed in Supplementary Table S2.
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After flowering, all floral buds of an inflorescence from the Ogura-CMS and MF lines of B. rapa ssp. rapifera were collected. In each case, samples were harvested and pooled from ten individual plants with transcriptome profiles representing ‘f’ difference, then immediately frozen in liquid nitrogen and stored at -70°C until RNA isolation. For biological repetitions, RNA was extracted from three samples using the EASYspin Plant RNA kit (Aidlab Biotechnologies Corporation, Bejing, China). RNA quality and quantity were characterized on a 1% agarose gel, and determined with a NanoPhotometer spectrophotometer (IMPLEN, CA, USA) and a Qubit RNA Assay Kit in Qubit2.0 Flurometer (Life Technologies, CA, USA). RNA integrity was assessed using the Agilent Bioanalyzer 2100 system (Agilent Technologies, CA, USA).
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The cotyledons and flowers tissues were collected from red cultivar ‘HHTQ’ and white cultivar ‘Beizaosheng’. The total RNA was extracted from the samples using EASYspin Plant RNA Kit (Aidlab, Wuhan, China) according to the manufacturer’s instructions, and were then treated with DNase I to remove genomic DNA [64 (link)]. RNA quality was checked and quantified using a Nanodrop 2000r (Nanodrop Technologies, Wilmington, DE, USA) and Agilent bioanalyzer 2100 (Agilent Technologies, Santa Clara, CA, USA), respectively.
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Total RNA from various plants was extracted using the EASYspin Plant RNA Kit (Aidlab, Beijing, China). Elimination of genomic DNA and the synthesis of first-strand cDNA were performed using the PrimeScript™ RT reagent Kit with gDNA Eraser (Takara, Dalian, China). qRT-PCR was conducted using SYBR® Premix Ex TaqTM II (Takara Bio Inc., Dalian, China), and the PCR program was as follows: one cycle of 3 min at 95 °C, followed by 40 cycles of 10 s at 95 °C and 30 s at 60 °C. Three genes, FtH3, AtActin2 and Ntβ-actin were used as an internal control for Tartary buckwheat, Arabidopsis and tobacco, respectively. The relative expression level of these genes in this study was calculated using the Change 2(−ΔΔCt) method. All analyses were conducted with three biological replicates for each sample. Primer sequences used for qRT-PCR are listed in Table S2.
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Total RNA of citrus leaves was extracted using an EASY Spin Plant RNA Kit according to the manufacturer’s instructions (Aidlab, China). cDNA was synthesized using 0.5–1.0 µg extracted RNA with a RevertAid First-Strand cDNA Synthesis Kit (Fermentas, Canada). Then, qPCR was carried out using iTaqTM Universal SYBR Green Supermix (Bio-Rad, USA) in a CFX96TM Real-Time System. A previously described thermal cycling program was used35 (link). Data were collected and analyzed with CFX Manager 3.1 software (Bio-Rad). qPCRs were performed in triplicate from three independent biological replicates of each sample. Actin was used as the internal control. The gene-specific primers used in the qPCR are displayed in Table S5.
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Total RNA was extracted from collected tissue samples of G. hirsutum using the EASYspin Plant RNA kit (Aidlab Biotech, Beijing, China). The quantity of RNA obtained was determined using a Nanodrop ND 2000 (Nanodrop Technologies, Wilmington, DE), and cDNA was synthesized using the SuperScript™ III Reverse Transcriptase Kit (Invitrogen, Carlsbad, CA) according to the manufacturer's instructions. The qPCR measurement was conducted on an ABI7500 PCR System (Applied Biosystems, Carlsbad, CA). Actin (GenBank accession number: AY305733) was used as reference gene. Gene‐specific primers (Table 2) for two target genes, and reference gene were designed using Beacon Designer 7.9 (Premier Biosoft, Palo Alto). All samples were assayed in 20 μL reaction systems using the Talent qPCR PreMix kit (Tiangen Biotech Co. Ltd, Beijing, China) according to the manufacturer's instruction. The PCR cycling parameters were as follows: 95 °C for 3 min, followed by 40 cycles of 95 °C for 5 s and 60 °C for 32 s. Three technical replicates were done for each sample.
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Two high‐quality upland cotton cultivars (HY405 and ND13) and two normal‐quality upland cotton cultivars (CCRI8 and ND601) were grown in the field of Baoding, China, in 2014. For the cotton fibre samples, bolls were collected at 0, 5, 10, 15, 20, 25 and 30 DPA. Samples from different plants were pooled. Total RNA was extracted from these samples using the EASY spin Plant RNA kit (Aidlab, Beijing, China). The qualified RNA was used for sequencing analysis using TopHat v2.0 on a HiSeq 2500 platform at the Novogene Bioinformatics Institute, Beijing, China.
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Total RNAs of eggplant root samples were isolated through EASYspin Plant RNA kit (Aidlab, China). Four cDNA libraries with 3 biological replicates each were constructed from reverse transcription of total RNA and sequenced with the Illumina HiSeq2500 (Illumina, CA, USA). The raw data have been uploaded in the SRA database of NCBI, and the BioProject accession number is PRJNA908752. To ensure high quality sequencing, the raw reads were processed to clean reads using fastp 0.18.0. Gene expression levels for each cDNA library were normalized by fragment per kilobase of transcript per million mapped reads (FPKM). Differentially expressed genes (DEGs) were identified by the criteria of false discovery rate (FDR) < 0.05 and |log2 fold-change value| ≥1. Gene Ontology (GO) and KEGG pathway analyses of the DEGs were accomplished through Blast2GO 2.8 and BLASTX.
RNA-Seq results were verified by quantitative real-time PCR (qPCR). The cDNA templates used for qPCR were obtained through the reverse transcription of the remaining total RNA with the help of PrimeScript™ RT Master Mix kit (Takara, Japan). Five DEGs were randomly selected for qPCR experiments on CFX96 Touch instrument (Bio-Rad Laboratories, Hercules, CA) coupled with SYBR Green II PCR Master Mix kit (Takara, Japan), and the qPCR results were calculated using the ΔΔCt method (Schmittgen and Livak, 2008 (link)).
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RNA extraction was used EASYspin Plant RNA Kit (Aidlab Biotech, Beijing, China). RNA reverse transcription used TranScript® II All-in-One First-Strand cDNA Synthesis SuperMix for qPCR (One-Step gDNA Removal) from TransGen Biotech. 1000 ng RNA to reverse transcribe into cDNA, and diluted the cDNA four times for qRT-PCR. The qRT-PCR was performed by using the Gene Applied Biosystems® 7500 Fast and TransStart Top Green qPCR SuperMix (TransGen Biotech, Beijing, China). qRT–PCR was conducted in a 20 μl volume containing 1 μl of diluted cDNAs, 0.4 μl of the each primer, 0.4 µL of passive reference dye, 7.8ul H2O, and 10 µL of Top Green qPCR SuperMix under the following conditions: 95 ◦C for 300 s, followed by 40 cycles of 95 ◦C for 15 s, 58 ◦C for 20 s and 72 ◦C for 30 s. GhActin gene was used as an internal reference gene. The 2−△△Ct method was used to calculate relative expression levels [37 (link), 38 ].
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