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Fluorescence quantitative internal reference gene under heat stress in haizhou changshan mountain and primers and application thereof

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

Inactive Publication Date: 2019-01-25
NANJING FORESTRY UNIV
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  • 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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  • Fluorescence quantitative internal reference gene under heat stress in haizhou changshan mountain and primers and application thereof
  • Fluorescence quantitative internal reference gene under heat stress in haizhou changshan mountain and primers and application thereof
  • Fluorescence quantitative internal reference gene under heat stress in haizhou changshan mountain and primers and application thereof

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Embodiment 1

[0033] 1. Plant tissue total RNA extraction and cDNA synthesis

[0034] 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.

[0035] 2. Selection of internal reference genes and design of primers

[0036] 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 under heat injury stress of Haizhou Changshan Mountain, primers and applications thereof, wherein the internal reference geneis MDH, EF-1A, aP-2 and UBC-E2, wherein the gene sequence of the MDH gene is shown in SEQ ID NO:10, EF-The gene sequence of 1A gene is shown in SEQ ID NO.11, aP-2 gene is shown in SEQ ID NO:13, UBC-The gene sequence of E2 gene is shown in SEQ ID NO.12. The present application screens out candidate genes with relatively stable expression according to the genes disclosed in the prior art, its stability is evaluated by software, 4 stable internal reference genes suitable for expression under heat stress in Haizhou Changshan mountain were obtain, based on these genes, real-time fluorescence quantitative PCR primers were designed, which filled the situation the there were no internal reference genes under heat stress in Haizhou Changshan Mountain, and provided a powerful support for the accurate quantification of related functional genes under heat stress in Haizhou Changshan Mountain, and could improve the stability, repeatability and reliability of the research.

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. technical background [0002] 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 verify the ...

Claims

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Application Information

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Patent Type & Authority Applications(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
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