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

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

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

According to the results of previous experiments, Haizhou Changshan has very good salt-alkali tolerance and can be used for afforestation and restoration of saline-alkali land. However, there has been no report on the genetic research of Haizhou Changshan under salt stress.

Method used

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  • Fluorescence quantitative internal reference gene under salt stress in haizhou changshan and primers and application thereof
  • Fluorescence quantitative internal reference gene under salt stress in haizhou changshan and primers and application thereof
  • Fluorescence quantitative internal reference gene under salt stress in haizhou changshan and primers and application thereof

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Embodiment 1

[0029] 1. Plant tissue total RNA extraction and cDNA synthesis

[0030] 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.

[0031] 2. Selection of internal reference genes and design of primers

[0032] Query the main internal reference genes in other plants according to the literature, and finally design 17 internal reference genes required for the experiment, 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. NO.9), MDH (SEQ ID NO.10), EF-1A (S...

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Abstract

The invention discloses a fluorescent quantitative internal reference gene under salt stress of Haizhou Changshan Mountain and primers and applications thereof, wherein the internal reference gene isUBC-E2, RPL and MDH, wherein the gene sequence of MDH gene is shown in SEQ ID NO:10, the gene sequence of RPL gene is shown in SEQ ID NO:3, the gene sequence of UBC-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, a most stable internal reference gene suitable for expression under salt stress condition in Haizhou Changshan mountain is disclosed, Based on these genes, real-time fluorescence quantitative PCR primers were designed, whichfilled the situation the there were no internal reference genes under salt stress in Haizhou Changshan Mountain, and provided a powerful support for the precise quantification of related functional genes under salt 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 in particular relates to a fluorescent quantitative internal reference gene and its primer and application under the stress of Changshan salt in 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. , very few pests and diseases, strong adaptability. In addition, the flower shape is peculiar and beautiful, the flowering period is long, the fruit color is bright, and the fruit-hanging period is long in autumn and winter. It is an excellent autumn flower and fruit viewing tree species. According to previous experiments, Haizhou Changshan has very good salt-alkali tolerance and can be used for afforestation and restoration of saline-alkali land. However, there has been no report on the genetic research of Haizhou Changshan under salt st...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6895C12Q1/6851C12N15/11
CPCC12Q1/6851C12Q1/6895C12Q2600/13C12Q2600/166C12Q2531/113C12Q2545/101C12Q2563/107
Inventor 杨秀莲华雅洁岳远征施婷婷王良桂刘华歧
Owner NANJING FORESTRY UNIV
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