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Paeonia ostii reference gene under drought stress, and special primer and application thereof

An internal reference gene and drought stress technology, applied to Fengdan internal reference gene and its special primers and application fields under drought stress, can solve problems such as inconsistency, error, and unstable expression level, and achieve the effect of improving stability and reliability

Active Publication Date: 2020-10-02
YANGZHOU UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In daily experiments, the expression levels of traditionally used internal reference genes may be unstable under different conditions, which may easily lead to wrong results or even contradictory conclusions

Method used

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  • Paeonia ostii reference gene under drought stress, and special primer and application thereof
  • Paeonia ostii reference gene under drought stress, and special primer and application thereof
  • Paeonia ostii reference gene under drought stress, and special primer and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0054] Example 1: Screening of Fengdan real-time fluorescent quantitative PCR internal reference genes under drought stress

[0055] Based on the RNA-seq database obtained from the leaves of Fengdan normal growth and drought stress for 12 days, the RPKM values ​​of 78,392 Unigenes were obtained, and 10 Unigenes with stable expression were found as candidate internal reference genes (see Table 1).

[0056] Table 1 RPKM values ​​representing the expression levels of each Unigene in RNA-seq

[0057] sequence number in transcriptome normal growth Drought stress for 12 days serial number Unigene0040750 9.53 9.69 SEQ ID NO.1 Unigene0029456 13.58 14.29 SEQ ID NO.2 Unigene0043536 19.05 18.33 SEQ ID NO.3 Unigene0041281 56.98 54.97 SEQ ID NO.4 Unigene0043455 9.08 8.90 SEQ ID NO.5 Unigene0041583 144.36 136.35 SEQ ID NO.6 Unigene0030732 5.34 5.62 SEQ ID NO.7 Unigene0032356 16.17 15.81 SEQ ID NO.8 U...

Embodiment 2

[0061] Example 2: Analysis of expression stability of candidate internal reference genes

[0062] Analysis of the expression stability of candidate internal reference genes in the leaves of Fengdan under different drought stress times: taking Fengdan three-year-old potted seedlings as plant materials, watering at 17:00 every day for 3 consecutive days before natural drought treatment, and then Natural drought treatments (0 days, 4 days, 8 days and 12 days) were carried out, and three biological replicates were set up for each treatment. After harvesting, they were quick-frozen with liquid nitrogen and stored at -80°C.

[0063] The sample was extracted with the MiniBEST Plant RNAExtraction Kit (Takara, Japan) kit, and the RNA concentration and OD value were detected using a nucleic acid protein analyzer, and the integrity of the total RNA was detected by 1.5% agarose gel electrophoresis; 5×M-MLV Buffer reverse transcriptase (TaKaRa, Japan) was used to synthesize cDNA using Olig...

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Abstract

The invention relates to a paeonia ostii reference gene under drought stress, and a special primer and application thereof. The reference gene is a TATA box binding protein TBP gene, an actin ACT1 gene, an actin ACT2 gene, a glyceraldehyde-3-phosphate dehydrogenase GAPDH gene, an eukaryote translation initiation factor eIF1 gene, a eukaryote translation initiation factor eIF2 gene, a tubulin alpha-TUB gene, a tubulin beta-TUB gene, an RNA polymerase II RNA Pol II gene or an RNA polymerase II transcription factor RP II gene. The nucleotide sequence of the gene is disclosed as the sequence table. The invention aims to provide a gene which can be stably expressed in the paeonia ostii gene expression profile under drought stress, and the gene is used as a paeonia ostii drought stress referencegene.

Description

technical field [0001] The invention belongs to the field of plant biotechnology, and in particular relates to an internal reference gene of Fengdan under drought stress, a special primer and application thereof. Background technique [0002] The internal reference gene is a gene whose expression level is not affected by the research conditions and is constantly expressed, and is often used for the correction of real-time fluorescent quantitative PCR detection. In daily experiments, the expression levels of traditionally used internal reference genes may be unstable under different conditions, which may easily lead to erroneous results or even contradictory conclusions. Therefore, it is particularly important to select and screen a suitable gene as an internal reference gene for reference and correction of gene expression. [0003] Fengdan (Paeonia ostii) is a perennial woody plant of Paeoniaceae Paeoniaceae, which is a variety of Yangshan peony. Because of its high medici...

Claims

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Application Information

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IPC IPC(8): C12N15/29C12N15/53C12Q1/6895C12Q1/6851C12N15/11
CPCC07K14/415C12N9/0008C12Q1/6851C12Q1/6895C12Q2600/166C12Q2545/101C12Q2563/107C12Q2531/113
Inventor 赵大球陶俊许聪栾雨婷
Owner YANGZHOU UNIV
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