The tea leaves were divided into two parts along the main vein of the leaves. Half of them were used to extract gDNA (RNA free) using a Plant Genomic DNA Isolation Kit (Tsingke, Beijing, China). The total RNA was extracted from the other half of the samples using a Plant Total RNA Isolation Kit (Vazyme, Nanjing, China). A total of 1% agarose gel electrophoresis was used to check the integrity of total RNA. The concentration of total RNA was measured with a NanoDrop ND 1000 spectrophotometer (Thermo Fisher Scientific, Waltham, MA, USA). A total of 1–2 μg of the total RNA was used to synthesize the first-strand complementary DNA (cDNA) with HiScript Q RT SuperMix for qPCR (+gDNA wiper) (Vazyme, Nanjing, China). Specific primers were used to amplify genes containing RNA editing sites using the gDNA and cDNA as templates, respectively (Table S1 ).
Plant genomic dna isolation kit
Manufactured by Tsingke
Sourced in China
The Plant Genomic DNA Isolation Kit is a laboratory product designed to efficiently extract and purify high-quality genomic DNA from plant samples. The kit utilizes a simple and effective protocol to isolate DNA that can be used for various downstream applications, such as PCR, sequencing, and molecular analysis.
Automatically generated - may contain errors
4 protocols using plant genomic dna isolation kit
1
RNA Extraction and cDNA Synthesis
2
Sequencing Rap Homologs in Transgenic Plants
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Genomic DNA was extracted from the rapCR transgenic plants and the wild‐type ‘Ningyu’ by a Plant Genomic DNA Isolation Kit (TSINGKE Biological Technology, Beijing, China, Cat# TSP101‐200). Full length sequences of the RAP homologs were amplified by PCR and inserted into the T vector by using the pEASY‐T1 Cloning Kit (TransGen Biotech, Beijing, China, Cat# CT101). A total of 10‐20 bacterial colonies for each transgenic plant were selected for Sanger sequencing to examine the induced mutations.
3
Genomic DNA Extraction and Methylation Analysis
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Genomic DNA was extracted from the woodland strawberry or Arabidopsis tissues using a plant genomic DNA isolation Kit (TSINGKE Biological Technology; cat no. TSP101-200). For strawberry samples, 100-ng genomic DNA was cleaved by the methylation sensitive enzyme McrBC for overnight, then the digested DNA was used as template to amplify the target sequences by qPCR, and the fragment in GAPDH (FvH4_4g24420) without DNA methylation was used as the control. For Arabidopsis samples, 100-ng genomic DNA was cleaved by HaeIII or McrBC for overnight, then the digested DNAs and the same amount of undigested DNAs were used for qPCR amplification at the target loci.
4
Targeted Sequencing of FveLFY Homologs
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Genomic DNA was extracted using a Plant Genomic DNA Isolation Kit (TSINGKE Biological Technology, Beijing, China). Full-length sequences of the FveLFY homologs were amplified by PCR. To determine the potential gene editing events, the PCR products were subjected to Sanger sequencing. In case of 2 different alleles exist in 1 plant, the PCR products were inserted into a T vector using a pEASY-T1 Cloning Kit (TransGen Biotech, Beijing, China) . A total of 10 bacterial colonies for each transgenic plant were sequenced. For each of the identified fvelfy CR mutants in this study, all the similar target sequences in their homologs were examined to exclude potential off-target effects.
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