Citrus total RNA was isolated using the EASYspin plant RNA extraction kit following the manufacturer’s instructions (Aidlab, Beijing, China). RNA was reverse transcribed into cDNA using the Prime ScriptRT Master Mix (TaKaRa, Ojin, Japan). RT-qPCR reaction system is 12 µL, including 6 µL the SYBRPRIME qPCR Kit (Bioround Biotechnology, Chongqing, China), 0.3 µL forward and reverse real-time primer (10 mM·L−1), 4.4 µL ddH2O and 1 µL cDNA (10 ng·µL−1). The PCR amplification conditions were: treatment at 95 °C for 2 min, then 40 amplification cycles (each at 65 °C for 10 s, 95 °C for 5 s), and finally extension at 60 °C for 15 s. Using citrus GAPDH gene (Mafra et al., 2012 (link)) as internal reference gene, the relative expression of mSDE460 in transgenic plants was calculated by the 2−ΔΔCt method (Livak and Schmittgen, 2001 (link)). The primers were listed in Supplementary Table
EASYspin plant RNA extraction kit
The EASYspin plant RNA extraction kit is a laboratory equipment designed for the efficient extraction and purification of high-quality total RNA from a wide range of plant tissues. It utilizes a simple and rapid spin-column procedure to isolate RNA, ensuring reliable and consistent results.
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29 protocols using EASYspin plant RNA extraction kit
Citrus RNA Extraction and RT-qPCR Analysis
Citrus total RNA was isolated using the EASYspin plant RNA extraction kit following the manufacturer’s instructions (Aidlab, Beijing, China). RNA was reverse transcribed into cDNA using the Prime ScriptRT Master Mix (TaKaRa, Ojin, Japan). RT-qPCR reaction system is 12 µL, including 6 µL the SYBRPRIME qPCR Kit (Bioround Biotechnology, Chongqing, China), 0.3 µL forward and reverse real-time primer (10 mM·L−1), 4.4 µL ddH2O and 1 µL cDNA (10 ng·µL−1). The PCR amplification conditions were: treatment at 95 °C for 2 min, then 40 amplification cycles (each at 65 °C for 10 s, 95 °C for 5 s), and finally extension at 60 °C for 15 s. Using citrus GAPDH gene (Mafra et al., 2012 (link)) as internal reference gene, the relative expression of mSDE460 in transgenic plants was calculated by the 2−ΔΔCt method (Livak and Schmittgen, 2001 (link)). The primers were listed in Supplementary Table
Extracting High-Quality RNA from Plant Leaves
Extraction and Analysis of Plant RNA
Quantification of gene expression via qRT-PCR
Heat and Drought Stress Response in R. chinensis
Quantifying CsSAMT1 Expression in Citrus
Total RNA Extraction and cDNA Synthesis
RNA Extraction and qRT-PCR Analysis
Quantitative RT-PCR Analysis of Plant Transcripts
RNA-Seq Analysis of Petal Development
Differential expression analysis of two conditions/groups was performed using the DESeq2 (Love et al. 2014 (link)). The resulting P-values were adjusted using the Benjamini and Hochberg’s approach for controlling the false discovery rate. Genes with an adjusted P-value < 0.05 (log2FoldChange > 2 | log2FoldChange < -2) found by DESeq2 were assigned as differentially expressed. The Venn diagram and kyoto encyclopedia of genes and genomes (KEGG) enrichment diagram were performed using R software.
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