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The internal reference gene of coral dish and its screening method and application

A technology of internal reference genes and screening methods, which is applied in the field of plant genetic engineering, can solve the problems of internal reference gene screening and stability verification without coral dish, and achieve the effects of improving scientific research work efficiency, improving calibration capabilities, and reducing costs

Active Publication Date: 2021-01-29
INST OF BOTANY JIANGSU PROVINCE & CHINESE ACADEMY OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0006] At present, there is no research report on the development, screening and stability verification of the internal reference gene of coral vegetable. In addition to the above reasons, the development of the stable internal reference gene of coral vegetable and its primers is of great importance in the molecular biology research of coral vegetable and umbrella. It has important practical application value in the research of plants

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  • The internal reference gene of coral dish and its screening method and application
  • The internal reference gene of coral dish and its screening method and application
  • The internal reference gene of coral dish and its screening method and application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0056] This example is an internal reference gene of the coral dish of the present invention. The following 13 internal reference genes of Corydalis are provided: PP2A (protein phosphatase 2), UBQ10 (ubiquitin 10), ACT (actin), EF1-α (elongation factor), GAPDH (glyceraldehyde-3-phosphate hydrogenase), α-TUB (α-tubulin), β-TUB (β-tubulin), PTBP1 (polypyrimidine sequence binding protein), EXP1 (expansin 1), EXP2 (expansin 2), TIP41-like (TIP41-like protein), SAND family (sand family protein), CYP2 (cyclophilin); the nucleotide sequence of the PP2A is SEQ ID NO.1; the nucleotide sequence of the UBQ10 is SEQ ID NO.2; the nucleotide sequence of the ACT is SEQ ID NO.3; the nucleotide sequence of the EF1-α is SEQ ID NO.4; the nucleotide sequence of the GAPDH is SEQ ID NO.5; The nucleotide sequence of the α-TUB is SEQ ID NO.6; the nucleotide sequence of the β-TUB is SEQ ID NO.7; the nucleotide sequence of the PTBP1 is SEQ ID NO.8; the nucleotide sequence of the β-TUB is SEQ ID NO.8; ...

Embodiment 2

[0058] In this example, fluorescent quantitative PCR primers for amplifying the internal reference gene in Example 1 are provided,

[0059] The size of the amplified fragment of the primers is 100-250bp.

[0060] The primer pair sequence used to amplify PP2A is SEQ ID NO.15 / SEQ ID NO.16; the primer pair sequence used to amplify UBQ10 is SEQ ID NO.17 / SEQ ID NO.18; the primer pair sequence used to amplify ACT The primer pair sequence is SEQ ID NO.19 / SEQID NO.20; the primer pair sequence for amplifying EF1-α is SEQ ID NO.21 / SEQ ID NO.22; the primer pair sequence for amplifying GAPDH is SEQ ID NO.21 / SEQ ID NO.22; ID NO.23 / SEQ ID NO.24; The primer pair sequence for amplifying α-TUB is SEQ ID NO.25 / SEQ ID NO.26; The primer pair sequence for amplifying β-TUB is SEQ ID NO. 27 / SEQ ID NO.28; the primer pair sequence used to amplify PTBP1 is SEQ ID NO.29 / SEQ ID NO.30; the primer pair sequence used to amplify EXP1 is SEQ ID NO.31 / SEQ ID NO. 32; the primer pair sequence used to amplify E...

Embodiment 3

[0062] The present embodiment provides a real-time fluorescent quantitative PCR method for screening the internal reference genes of the above-mentioned Anchovy, which comprises the following steps: (1) selecting root tissues of Anchophylla that are respectively treated with salt stress, drought stress, abscisic acid, and methyl jasmonate sample;

[0063] (2) Use the sequencing data of the transcriptome of the coral dish to screen out the candidate internal reference genes of the coral dish;

[0064] (3) Use the selected candidate internal reference genes as templates (i.e. PP2A, UBQ10, ACT, EF1-α, GAPDH, α-TUB, β-TUB, PTBP1, EXP1, EXP2, TIP41-like, SAND family, CYP2; specific For the sequence, please refer to the sequence table (SEQ ID NO.1-SEQ ID NO.13), design and obtain the primers for real-time fluorescent quantitative PCR of each internal reference gene (see Example 2 for details);

[0065] (4) Perform real-time fluorescent quantitative PCR, perform statistical analysis...

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Abstract

The invention discloses an internal reference gene of coral dish and its screening method and application, and belongs to the technical field of plant genetic engineering. It specifically relates to an internal reference gene and primers thereof of coral dish, said internal reference gene is PP2A, UBQ10, ACT, EF1-α, GAPDH, α-TUB, β-TUB, PTBP1, EXP1, EXP2, TIP41-like, SAND family, CYP2 and provide a real-time fluorescent quantitative PCR screening method for internal reference genes of coral vegetables, which fills the blank of lacking suitable and general internal reference genes in the field of coral vegetable research.

Description

technical field [0001] The invention belongs to the technical field of plant genetic engineering, and more specifically relates to an internal reference gene of coral dish, its screening method and application. Background technique [0002] Coral vegetable is a herbaceous plant of the Umbelliferae family. Its roots are used as medicine. It is named Adenophora. It is a traditional Chinese medicinal material in my country. Lung, Stomach-invigorating and fluid-promoting effect, it is mainly used to treat diseases such as lung heat, yin deficiency, dry cough, dry stomach, dry throat and thirst. It is commonly used in clinical and health care, and the annual demand in domestic and foreign markets can reach more than 8,000 tons. [0003] There are two main distribution areas for the wild resources of coral vegetables: one is the distribution area of ​​coral vegetables in East Asia, and the other is the distribution area of ​​coral vegetables in North America. China is one of the ma...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/6895C12Q1/6851C12N15/11
CPCC12Q1/6895C12Q2600/166C12Q2600/158C12Q1/686C12Q2600/13C12Q2561/113C12Q2563/107C12Q2545/114
Inventor 李莉周义峰李乃伟
Owner INST OF BOTANY JIANGSU PROVINCE & CHINESE ACADEMY OF SCI
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