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Fluorescent quantitative internal reference genes and their primers and applications under heat stress in Changshan, Haizhou

A fluorescence quantitative, internal reference gene technology, applied in the field of plant molecular biology, to achieve the effect of improving detection efficiency, strong primer specificity, and improving reliability

Inactive Publication Date: 2019-12-27
NANJING FORESTRY UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

There is no report on the gene research under heat stress in Changshan, Haizhou

Method used

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  • Fluorescent quantitative internal reference genes and their primers and applications under heat stress in Changshan, Haizhou
  • Fluorescent quantitative internal reference genes and their primers and applications under heat stress in Changshan, Haizhou
  • Fluorescent quantitative internal reference genes and their primers and applications under heat stress in Changshan, Haizhou

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Embodiment 1

[0034] 1. Plant tissue total RNA extraction and cDNA synthesis

[0035] Extract total RNA with EASYspin Plus Plant RNA Extraction Kit (Adley Biotechnology Co., Ltd.), use a spectrophotometer (METTLER TOLEDO, Switzerland) to detect RNA concentration and quality, and utilize 1.5% agarose gel electrophoresis to detect the total complete RNA. sex. Using Beijing Quanshijin Reverse Transcription Kit (Quanshijin Biotechnology Co., Ltd.), RNA was purified and the first strand of cDNA was synthesized according to the instructions.

[0036] 2. Selection of internal reference genes and design of primers

[0037] Query the main internal reference genes in other plants according to the literature and finally design 17 internal reference genes required by this embodiment, which are Actin (SEQ ID NO.1), PP2A (SEQ ID NO.2), RPL (SEQ ID NO. .3), PK (SEQ ID NO.4), 18S (SEQ ID NO.5), RAN (SEQ ID NO.6), APT (SEQ ID NO.7), SAND (SEQ ID NO.8), PROF ( SEQ ID NO.9), MDH (SEQ ID NO.10), EF-1A (SEQ ...

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Abstract

The invention discloses a fluorescent quantitative internal reference gene and its primers and applications under heat stress in Changshan, Haizhou. The internal reference genes are MDH, EF-1A, AP-2 and UBC-E2, wherein the gene sequence of the MDH gene is as SEQ ID NO As shown in .10, the gene sequence of the EF-1A gene is shown in SEQ ID NO.11, the gene sequence of the AP-2 gene is shown in SEQ ID NO.13, and the gene sequence of the UBC-E2 gene is shown in SEQ ID NO.12 shown. This application screens out candidate genes with relatively stable expression based on the existing published genes, evaluates their stability through software, and obtains 4 internal reference genes suitable for stable expression under heat stress conditions in Changshan, Haizhou, and uses these genes as the basis for design The real-time fluorescence quantitative PCR primers were introduced to fill in the current situation that there were no internal reference genes under the heat stress conditions in Changshan, Haizhou, and provide strong support for the accurate quantification of related functional genes under the heat stress conditions in Changshan, Haizhou, which can improve the stability and repeatability of research. sex and reliability.

Description

technical field [0001] The invention belongs to the field of plant molecular biology, and specifically relates to a fluorescent quantitative internal reference gene and its primer and application under heat stress in Changshan, Haizhou. [0002] technical background [0003] Haizhou Changshan (Clerodendrum trichotomum Thunb.), also known as stinky phoenix tree, is a deciduous shrub or small tree under the genus Clerodendmm L. of Verbenaceae. , also heat-resistant, very few pests and diseases, strong adaptability. In addition, the flower shape is peculiar and beautiful, the flowering period is long, and the fruit is brightly colored. It is an excellent tree species for viewing flowers and fruits in autumn. There is no report on the gene research under heat stress in Changshan, Haizhou. In the process of exploring the differences in gene expression under heat stress in Changshan, Haizhou, it is necessary to use real-time fluorescence quantitative technology to analyze and ver...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/6895C12Q1/686C12N15/11
CPCC12Q1/686C12Q1/6895C12Q2600/166C12Q2561/113C12Q2563/107C12Q2545/114C12Q2521/107
Inventor 杨秀莲刘华歧岳远征施婷婷王良桂华雅洁
Owner NANJING FORESTRY UNIV