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Primer composition, kit and method for compositely identifying polymorphic genetic marker of cannabis sativa

A primer composition and polymorphic inheritance technology are applied in the field of primer compositions for compound identification of polymorphic genetic markers in cannabis, and achieve the effects of strong stability, good specificity and high sensitivity

Pending Publication Date: 2022-04-22
ACADEMY OF FORENSIC SCIENCE
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The foreign research on the cannabis STR compound amplification system is in its infancy, and there is almost no research on it in China

Method used

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  • Primer composition, kit and method for compositely identifying polymorphic genetic marker of cannabis sativa
  • Primer composition, kit and method for compositely identifying polymorphic genetic marker of cannabis sativa
  • Primer composition, kit and method for compositely identifying polymorphic genetic marker of cannabis sativa

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0048] The present embodiment provides a method for polymorphic genetic markers for compound identification of marijuana, comprising the steps of:

[0049] 1. Screening of cannabis STR loci suitable for forensic applications;

[0050] According to literature reports and gene databases, search for the currently developed cannabis STR loci, but not all of these STR loci are suitable for the research of cannabis multiple amplification systems. Screen the polymorphic STR loci and eliminate the loci with dinucleotide repeats. Finally, 17 optimal autosomal STR loci and 2 sex identification loci were selected for kit development. The loci are D02-CANN1, C11-CANN1, DM029, DM016, 4910, B01-CANN1, E07- CANN1, 9269, B05-CANN1, H06-CANN2, 5159, nH09, ANUCS 501, CS1, ANUCS 305, 3735, ANUCS302, 1528 and 9043. Among them, the gender identification site DM029 is related to the X chromosome, and all samples have a single peak, and the sex identification site DM016 is related to the Y chromos...

Embodiment 2

[0068] In this example, the method provided in Example 1 is used to detect the DNA of a sample of the genus Cannabis.

[0069] Different tissues (flowers, stems, leaves, and seeds) of a Cannabis sample were collected for DNA extraction and quantification. The kit constructed in Example 1 was used to perform multiplex amplification of 19 genetic markers on the above DNA samples. All samples obtained effective amplification products on genetic markers.

Embodiment 3

[0071] According to the requirements of SWGDAM, this embodiment performs forensic verification (identity, sensitivity, species specificity and forensic parameters calculation) on the method provided in Example 1, and then judges the superiority of the method. The specific operations are as follows:

[0072] (1) Identity study: Quantify the genomic DNA of flowers, leaves and stems of the same plant to 1 ng / μL, and use the method provided in Example 1 to detect them respectively.

[0073] The results show( Figure 4 ) The kit constructed in Example 1 has good tissue identity: the flowers, leaves and stems of the same plant have the same genotype.

[0074] (2) Sensitivity study: the cannabis genomic DNA was diluted to 2ng / μL, 1ng / μL, 0.5ng / μL, 0.25ng / μL, 0.125ng / μL, 0.0625ng / μL, 0.03125ng / μL and 0.015625ng / μL μL, use the method provided in Example 1 to detect the cannabis genomic DNA at the above concentration, and repeat the detection 3 times for each template sample.

[0075]...

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Abstract

The invention provides a primer composition, a kit and a method capable of detecting 17 autosomal STR gene loci and two sex identification loci of a cannabis sample in parallel, the 17 autosomal STR gene loci comprise D02-CANN1, C11-CANN1, 4910, B01-CANN1, E07-CANN1, 9269, B05-CANN1, H06-CANN2, 5159, nH09, ANUCS 501, CS1, ANUCS 305, 3735, ANUCS 302, 152 and 9043, and the two sex identification loci are DM029 and DM016; the primer composition comprises the sequences as shown in SEQ ID NO. 1 to 38. The primer composition, the kit and the method provided by the invention have the characteristics of high sensitivity, good specificity, strong stability, high accuracy and the like, and can be used for detecting 17 autosomal STR gene loci and 2 sex identification loci of a cannabis sample in parallel. A brand new detection means is provided for sex identification, individual identification, source inference and STR database establishment in forensic medicine research of marihuana in China.

Description

technical field [0001] The invention relates to the technical field of molecular identification, in particular to a primer composition, a kit and a method for compound identification of polymorphic genetic markers in cannabis. Background technique [0002] Cannabis is an annual dioecious herbaceous plant belonging to the family Cannabinaceae (Cannabis). It is mainly distributed in Nepal, India, China and other regions in Europe, Africa and Asia. It is one of the oldest cultivated crops on the earth. Hemp fiber is a good raw material for textile and papermaking. The cannabinoids contained in the plant are widely used in the field of medicine and health. The seeds are rich in protein, amino acids, unsaturated fatty acids and trace elements that are easily absorbed by the human body. An important economic crop in many countries. However, due to the strong addictive and narcotic properties of tetrahydrocannabinol (THC) contained in the flowers and leaves, marijuana has been lis...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6895C12Q1/6879C12Q1/686C12N15/11
CPCC12Q1/6895C12Q1/6879C12Q1/686C12Q2600/156C12Q2525/151
Inventor 张素华李成涛夏若成于欢陶瑞旸陈安琪刘希玲阙庭志林源赵珍敏
Owner ACADEMY OF FORENSIC SCIENCE