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A kind of primer pair and test kit and application for identification of cigar tobacco

The technology of a kit and primer pair is applied in biochemical equipment and methods, microbiological measurement/testing, DNA/RNA fragments, etc., which can solve the problems of narrow genetic basis and unfavorable development of tobacco breeding, and achieve simple screening methods and improved Breeding process, simple operation effect

Active Publication Date: 2021-10-15
中国烟草总公司海南省公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Xia et al. used 28 pairs of SSR markers to analyze the genetic diversity and predict the population structure of 78 tobacco cultivars in my country. The results showed that the genetic basis of the 78 cultivars was relatively narrow, which was not conducive to the development of tobacco breeding.

Method used

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  • A kind of primer pair and test kit and application for identification of cigar tobacco
  • A kind of primer pair and test kit and application for identification of cigar tobacco
  • A kind of primer pair and test kit and application for identification of cigar tobacco

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0037] Using the medium-term bank of national germplasm resources of Tobacco Research Institute of Chinese Academy of Agricultural Sciences and Hainan Cigar Research Institute, 5 cultivars are commonly used in cigar producing countries: H211 (China), Beinhart 1000-1 (USA), Little Dutch (USA) , Havana 10 (Cuba), H382 (Indonesia) and Hicks (SSR marker positive control) and 20 cigar resources of different geographical origins (as shown in Table 2) carry out the screening of cigar tobacco SSR primer pairs:

[0038] Table 2. Cigar resource name and source

[0039]

[0040]

[0041] At the stage of 5 true leaves of the test sample resources, 10 plants were randomly selected for mixed sampling, genomic DNA was extracted by CTAB method, DNA was dissolved by TE, and RNase was added to water bath for 30 minutes. The quality and concentration of DNA were measured by a nucleic acid analyzer (NanoProp 2000 Thermoscientific), and the DNA concentration was uniformly diluted to 50ng / μL,...

Embodiment 2

[0057] Select PCR amplified bands with a clear band type for manual reading, and record bands as "1" and no bands as "0", establish a "0 / 1" matrix, and use DataFormater software to convert the matrix. Calculated with Popgene32 software: observed number of alleles Na (observed number of alleles), effective allele Ne (effective number of alleles), Nei's gene diversity index H (Nei's gene diversity), Shannon's polymorphic information index I (Shannon's information index) and the paired Nei's genetic distance (Nei's genetic distance) between varieties, the results are shown in Table 5:

[0058] Table 5. Polymorphism information of cigar tobacco SSR primers in the tested cigar tobacco resources

[0059]

[0060]

[0061]

[0062] The results showed that the genetic diversity of 48 pairs of SSR markers in 20 cigar tobacco resources showed that the number of observed alleles was between 2 and 9, with an average of 4.02, and the number of alleles in primer PT52689 was at most...

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Abstract

The invention provides a pair of primers, a kit and application for identification of cigar tobacco, and belongs to the field of DNA marker technology and application of molecular biology. The screening method of the present invention comprises the following steps: using tobacco genomic DNA as template DNA; using published public SSR markers as primers to be screened, performing PCR amplification on the primers to be screened and template DNA to obtain PCR products; The product is detected, and the target fragment can be observed as the SSR primer. The SSR core primers obtained by the present invention have the advantages of high polymorphism, good stability, easy operation and promotion, uniform distribution, etc., can be directly used, and greatly improve the screening efficiency. Breeding work provides strong technical support and is of great significance to cigar research.

Description

technical field [0001] The invention belongs to the field of DNA marker technology and application of molecular biology, and specifically relates to a primer pair, a kit and an application for identification of cigar tobacco. Background technique [0002] Cigars are powerful, full-bodied, rich in flavor, and have a small ratio of tar to nicotine. In recent years, the demand in domestic and foreign markets has been increasing, and the industry has broad prospects. my country's cigar tobacco research work started late, and the basic research is backward. Most of the varieties currently planted are foreign varieties. Restricted by planting conditions and cultivation measures, there is still a big gap between the quality of tobacco leaves and foreign high-quality cigar leaves, which cannot meet the needs of tobacco industry companies for tobacco leaf raw materials. Therefore, constructing cigar tobacco core primers to analyze cigar tobacco genetic relationship and genetic diver...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/6895C12N15/11
CPCC12Q1/6895C12Q2600/156
Inventor 刘国祥张兴伟吕洪坤梁开朝李媛向小华刘好宝戴培刚佟英杨爱国冯全福
Owner 中国烟草总公司海南省公司
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