Screening and application of high-expression reference gene Ce047468 of taro

An internal reference gene and high-expression technology, which is applied in the fields of application, genetic engineering, plant gene improvement, etc., can solve the problem of high expression level and achieve the effect of high expression level

Active Publication Date: 2021-12-28
GUANGXI ZHUANG AUTONOMOUS REGION ACAD OF AGRI SCI
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AI Technical Summary

Problems solved by technology

The reliability of qRT-PCR results is affected by factors such as initial sample size, RNA integrity, cDNA quality, and amplification efficiency. An internal reference gene with stable expression is crucial for comparing the expression levels of target genes in different samples. Correct selection of an appropriate internal reference gene Correction and standardization are required to obtain accurate qRT-PCR results. In September 2021, the applicant submitted a patent entitled "Internal Reference Gene Screening and Application in Taro Bulb Development", but the expression level of this application is not as good as this one. The application is high. For this reason, in order to obtain the reference genes of taro with higher expression levels during the development of taro bulbs, it is necessary to conduct more in-depth research

Method used

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  • Screening and application of high-expression reference gene Ce047468 of taro
  • Screening and application of high-expression reference gene Ce047468 of taro
  • Screening and application of high-expression reference gene Ce047468 of taro

Examples

Experimental program
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Effect test

Embodiment 1

[0028] This embodiment is the acquisition of the internal reference gene Ce047468, which is obtained by the following method:

[0029] One, the test material of present embodiment:

[0030] The taro material used in this example is Guiyu No. 2, a new taro variety bred by the Institute of Biotechnology, Guangxi Academy of Agricultural Sciences. In March 2020, it was planted in Guangxi Academy of Agricultural Sciences to build a scientific research base. Sampling was carried out continuously throughout the corm development period, and regular sampling was performed every 30 days. 30d, 60d, 90d, 120d, 150d, 180d, 210d, and 240d after sowing were collected from the corms, a total of 8 groups of samples were marked as YTS1, YTS2, YTS3, YTS4, YTS5, YTS6, YTS7, and YTS8. Each group of samples had three biological replicates. Peel the collected bulbs, cut them into small pieces of about 2 cm, put them into 50 ml sterile cryovials, freeze them with liquid nitrogen and put them in a ...

Embodiment 2

[0074]The gene Ce047468 with the best stability screened out in Example 1 was further verified to ensure that true and reliable quantitative data can be obtained. Taking Ce047468 as an internal reference gene, a gene related to bulb starch synthesis, the large subunit gene CeAGPL1 of ADP-glucose pyrophosphorylase (AGPase), was selected to verify the stability of the screened internal reference gene . Using Ce047468 as the internal reference gene and CeAGPL1 as the target gene to design primers (Table 5), qRT-PCR expression analysis was performed on the CeAGPL1 gene in eight developmental stages (YTS1-YTS8) of taro bulbs. The results showed that the internal reference gene was stably expressed and could It is better to reflect the expression level changes of CeAGPS1 in the corm development process, as follows:

[0075] Different stages were extracted respectively: that is, the entire development period of taro, that is, the bulbs of 30d, 60d, 90d, 120d, 150d, 180d, 210d, and 2...

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Abstract

The invention relates to the technical field of plant genes, in particular to screening and application of a high-expression reference gene Ce047468 of taro, and compared with a reference gene applied in a patent submitted by the applicant in September 2021 and named as reference gene screening in taro corm development process and application thereof, the reference gene Ce047468 has the advantages that the expression quantity is higher and more stable; according to the reference gene analysis mode, continuous sampling is carried out on the taro corm in the whole development period, regular sampling is carried out every 30 days, the taro corm material is analyzed according to RNA-Seq sequencing data, and the reference gene Ce047468 which is more stable than a common housekeeping gene is screened out from a high expression area. And a solid foundation is laid for exploring key genes related to colocasia esculenta corm development by using q RT-PCR in the later period.

Description

【Technical field】 [0001] The invention relates to the technical field of plant genes, in particular to the screening and application of the highly expressed internal reference gene Ce047468 of Taro. 【Background technique】 [0002] The edible part of taro is corm, which is rich in nutrients. The quality and output of taro in Guangxi are among the top in the country, especially Lipu taro in Guangxi. Therefore, taro has abundant germplasm resources in Guangxi. However, Because taro is not a major agricultural product, there are few studies on molecular biology, and there are few reports on its gene research. Real-time fluorescent quantitative PCR (quantitative Real-time PCR, qRT-PCR) is currently the main analytical method for detecting gene expression levels. The reliability of qRT-PCR results is affected by factors such as initial sample size, RNA integrity, cDNA quality, and amplification efficiency. An internal reference gene with stable expression is crucial for comparing...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/29C12Q1/6895C12Q1/6851C12N15/11
CPCC07K14/415C12Q1/6895C12Q1/6851C12Q2600/166C12Q2600/13C12Q2531/113C12Q2545/101C12Q2563/107
Inventor 董伟清何芳练刘莉莉胡祥红蒋慧萍邱祖杨
Owner GUANGXI ZHUANG AUTONOMOUS REGION ACAD OF AGRI SCI
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