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SNP markers for authenticating tortoises, Chinese stripe-necked turtles and hybrids of tortoises and Chinese stripe-necked turtles

A hybrid and marking technology, which is applied in the determination/inspection of microorganisms, biochemical equipment and methods, etc., can solve the problems of adulterated products mixed into the market, similar morphological characteristics, and difficult identification of three turtles, etc., to achieve The effect of accurate and reliable detection means and efficient detection means

Active Publication Date: 2019-10-11
湖北省药品监督检验研究院
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Due to the close relationship between the flower tortoise and the tortoise, their hybrids take into account the traits of the parents, which brings difficulties to the identification of the three tortoises
The current traditional identification method is mainly based on the external characteristics of turtles. This method is highly dependent on experience and has a low accuracy rate. Misjudgments are often made due to lack of experience, similar morphological characteristics or no obvious morphological characteristics of turtle products. Cause counterfeit or adulterated products of tortoise shells to mix into the market, bringing adverse consequences
[0003] In recent years, molecular identification technology has been widely used in the identification of traditional Chinese medicinal materials due to its advantages of rapidity and accuracy. distinguish it

Method used

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  • SNP markers for authenticating tortoises, Chinese stripe-necked turtles and hybrids of tortoises and Chinese stripe-necked turtles
  • SNP markers for authenticating tortoises, Chinese stripe-necked turtles and hybrids of tortoises and Chinese stripe-necked turtles
  • SNP markers for authenticating tortoises, Chinese stripe-necked turtles and hybrids of tortoises and Chinese stripe-necked turtles

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0032] Example 1. Obtaining of SNP molecular markers

[0033] The applicant used the GBS (Genotyping-by-Sequencing) sequencing technology to perform simplified genome sequencing on the flower tortoise sample, obtained the simplified genome sequence information of the flower tortoise, and used the whole genome information of the tortoise as a reference. The reads obtained by simplified genome sequencing were filtered out low-depth and low-integrity sequences, and compared with the tortoise genome information to obtain SNP molecular markers, these SNP sites were compared and observed to find stable ones suitable for species identification The SNP site is found to specifically exist in flower tortoises, tortoises and their hybrids, and the SNP detection rate of the three turtles is guaranteed to be 100%, the frequency of base mutations to occur is 100%, and the upstream and downstream sequences of the mutation sites are highly conserved Finally, four groups of SNP molecular marke...

Embodiment 2

[0046] Example 2. Gene source identification of tortoises, flower tortoises and their hybrids based on SNP sites

[0047] The screened SNP sites were compared to the reference genome for positioning, and a sequence including the SNP was obtained. It was found that there was a highly conserved region upstream and downstream of the SNP sites of the three turtles, and then a design was designed to identify the three turtles. The primer pair, the primer pair sequence is shown in Table 2.

[0048] Table 2 Primer pairs for amplifying four groups of sequences

[0049]

[0050] The designed primer pair is composed of upstream and downstream primer sequences, which are sent to Sangon Bioengineering Co., Ltd. for synthesis. Each primer is individually packaged and stored, and is added to the PCR primer reaction system in the form of primer solution and stored at -20°C. , the concentration of the primer solution was 10 pmol / μL.

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Abstract

The invention discloses SNP markers for authenticating tortoises, Chinese stripe-necked turtles and hybrids of the tortoises and the Chinese stripe-necked turtles. The SNP markers include the first SNP marker (a basic group A or G located on the 88th site of the nucleotide sequence shown in SEQ ID NO:1), the second SNP marker (a basic group T or C located on the 105th site of the nucleotide sequence shown in SEQ ID NO:2), the third SNP marker (a basic group C or T located on the 178th site of the nucleotide sequence shown in SEQ ID NO:3), and the fourth SNP marker (a basic group C or T locatedon the 233rd site of the nucleotide sequence shown in SEQ ID NO:4). Primers are designed and a detecting method is established for the markers, the problem that it is difficult to authenticate the tortoises, the Chinese stripe-necked turtles and the hybrids of the tortoises and the Chinese stripe-necked turtle is solved, and the accurate, reliable and efficient detecting means is provided for authenticating and supervising the authenticity of tortoise shell medicinal materials.

Description

technical field [0001] The present invention relates to SNP markers for identifying tortoises, flower tortoises and hybrids thereof, and primer pairs for detecting said SNP markers. The use in hybrids belongs to the field of molecular identification. Background technique [0002] Turtle (Chincmys reevesii) is a traditional animal medicine for both medicine and food in my country. "Shen Nong's Materia Medica" lists tortoise shell as the top grade, and "Compendium of Materia Medica" clearly describes the medicinal value of tortoise shell, turtle meat, and turtle blood. The national standard contains nearly a hundred prescriptions for tortoise shells. Some classic products related to tortoises, such as tortoise shell glue and Guilu Erxian ointment, are well-known at home and abroad, and are widely used in clinical treatment and nourishing health. Tortoise is the legal source of tortoise shell, a medicinal material included in the Pharmacopoeia of the People's Republic of China...

Claims

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Application Information

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IPC IPC(8): C12Q1/6888C12Q1/6869
CPCC12Q1/6888C12Q1/6869C12Q2600/156C12Q2600/124C12Q2531/113
Inventor 汪波聂晶胡敏徐玲肖凌周豫新黄小清覃桂
Owner 湖北省药品监督检验研究院
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