Passiflora internal reference gene penadp and its screening method and application
An internal reference gene, the technology of passionflower, applied in the field of molecular biology, can solve the problems that restrict the research process of passionflower cloning, etc., and achieve the effects of shortening the detection time, improving the reliability, and optimizing the PCR amplification program
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Embodiment 1
[0031] Passiflora internal reference gene PeNADP, the nucleotide sequence of the internal reference gene PeNADP is shown in SEQ ID NO.1.
Embodiment 2
[0033] A primer for PCR amplification of the internal reference gene PeNADP, nucleotide sequence: the forward sequence is shown in SEQ ID NO.2; the reverse sequence is shown in SEQ ID NO.3.
Embodiment 3
[0035] A screening method for Passiflora internal reference gene PeNADP, comprising the following steps:
[0036] S1. Download 4 sample data from the NCBI database, the sequence accession numbers are: SRX4224487, SRX4224486, SRX4224485 and SRX4224484;
[0037] Use Trinity (version r20131110) software, parameter min_kmer_cov 2 to assemble the sequence;
[0038] Use RSEM (v1.2.3) to analyze the expression level of predicted sequences;
[0039] Functional annotation of gene sequences using BLAST (v2.2.31) and databases NR, COG, KOG, eggNOG, KEGG, GO, Pfam and Swiss-prot;
[0040] The stability of the expression value of the PeNADP gene in the four samples was calculated, and the coefficient of variation (CV) was used for evaluation. The genes with a CV value less than 0.2 and cDNA ≥ 1000 bp were selected as candidate memory genes.
[0041] S2. Among the candidate genes, according to the expression value of PeNADP, 2 genes with the highest expression and 2 genes with medium and ...
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