PCR primer set, site, method and kit for minimally invasive and rapid identification of ZW sex chromosome type of pelodiscus sinensis

A Chinese soft-shelled turtle and chromosome technology, applied in biochemical equipment and methods, microbial determination/inspection, recombinant DNA technology, etc., can solve the problems of cumbersome operation, trauma of the individual to be inspected, sacrifice of the individual to be inspected, etc., and achieve accurate and intuitive results. , the effect of easy operation

Active Publication Date: 2019-04-19
ZHEJIANG FISHERIES TECH EXTENSION STATION
View PDF3 Cites 5 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, these methods are cumbersome to operate, and need to sacrifice the individual to be tested or cause

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • PCR primer set, site, method and kit for minimally invasive and rapid identification of ZW sex chromosome type of pelodiscus sinensis
  • PCR primer set, site, method and kit for minimally invasive and rapid identification of ZW sex chromosome type of pelodiscus sinensis
  • PCR primer set, site, method and kit for minimally invasive and rapid identification of ZW sex chromosome type of pelodiscus sinensis

Examples

Experimental program
Comparison scheme
Effect test

Example Embodiment

[0041] Example 1

[0042] The primers provided in this example for quickly identifying the ZW sex chromotype of the Chinese soft-shelled turtle include 1 forward primer and 1 reverse primer, wherein:

[0043] The ZW sex chromosome identification forward primer of Chinese soft-shelled turtle is shown in SEQ ID No.1;

[0044] The reverse primer for ZW sex karyotype identification of Chinese soft-shelled turtle is shown in SEQ ID No.2.

[0045] specific:

[0046] The forward primer sequence is: 5’-TGGTGCTGTRAAGAAGGMRRCT-3’,

[0047] The reverse primer sequence is: 5’-TTCTTCATCTTCCTCCTCRGGT-3’

[0048] This method is based on the ZW sex chromosome-linked gene sequence database obtained from the comparative analysis of NCBI Chinese soft-shelled turtle genome (Pelsin-1.0). According to the difference fragments, a pair of Chinese soft-shelled turtle ZW sex chromotype identification primers were designed and verified.

Example Embodiment

[0049] Example 2

[0050] The PCR method for quickly identifying the ZW sex chromosome of the Chinese soft-shelled turtle provided in this embodiment includes the following steps:

[0051] (1) Sampling of Chinese soft-shelled turtle tissue to be inspected

[0052] Randomly select 10 male and female adult Chinese soft-shelled turtles that have been judged by appearance (samples 1-5 are female soft-shelled soft-shelled turtles from Japan; sample 6-10 are female soft-shelled turtles from Qingxi black soft-shelled turtle; samples 11-15 are from Japanese soft-shelled turtles Male soft-shelled turtle; No. 16-20 sample is Qingxi black soft-shelled turtle male). After the gonads are dissected to verify the physiological sex, a sample of hind limb muscle tissue is cut and stored at -80°C after flash freezing in liquid nitrogen for later use;

[0053] (2) Genomic DNA extraction

[0054] a) Using blood / cell / tissue genomic DNA extraction kit (Tiangen DP304). Take 30 mg of muscle tissue, add 200 μ...

Example Embodiment

[0072] Example 3

[0073] The PCR method for rapid and minimally invasive identification of the ZW sex chromosome of the Chinese soft-shelled turtle provided in this embodiment includes the following steps: (1) Blood sample collection of the newly hatched soft-shelled turtle

[0074] Randomly select 10 juvenile turtles from Qingxi Black Turtle just out of their shells, and use a syringe needle to open a hole in the yolk sac of the 1-5 juveniles. Use a 10 μL pipette to collect 1 μL of blood sample from the yolk sac and transfer it to the PCR tube; 6 No.-10 only uses a syringe needle to puncture the skin of the hind limbs, collect 1μL of peripheral blood with a 10μL pipette, and transfer it to the PCR tube. After blood collection, each juvenile turtle was placed in a separate water basin, treated with 10ppm potassium permanganate for 5 minutes, and kept under observation for five days. There was no wound infection, ulceration and death. The sex of the juvenile turtle was identified a...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

PUM

No PUM Login to view more

Abstract

The invention discloses a PCR primer set, site, method and kit for minimally invasive and rapid identification of the ZW sex chromosome type of pelodiscus sinensis. The sequence of the primer set is shown as SEQ ID No.1 and SEQ ID No.2. With a trace amount (0.5-1 microliter) of pelodiscus sinensis blood as a starter template, the primer set is used for carrying out PCR amplification, an amplification product is subjected to electrophoresis detection, and when 2031 bp bands appear in the amplification product, it is determined that the pelodiscus sinensis is in a ZZ sex chromosome type; when 2031 bp and 1624 bp bands appear in the amplification product, it is determined that the pelodiscus sinensis is in a ZW sex chromosome type. With the trace amount of blood obtained by minimally invasivecollection as a starter material, the sex chromosome type of the pelodiscus sinensis is rapidly detected; the detection hardly damages tested individuals, it can be ensured that pelodiscus sinensis in each growth and development stage can survive healthily after sampling, and an important tool is provided for identifying the sex hromosome type in the pelodiscus sinensis monosex culture technologyand sex-controlled breeding.

Description

technical field [0001] The invention belongs to the technical field of molecular biology identification, relates to the field of identification of the sex chromosome type of soft-shelled soft-shelled turtle ZZ / ZW, and in particular relates to a PCR amplification primer set and a positioner for minimally invasive and rapid identification of the sex chromosome type of soft-shelled soft-shelled turtle, which can save the genome extraction operation. Points, methods and kits. Background technique [0002] The Chinese soft-shelled soft-shelled turtle (Pelodiscus sinensis) belongs to the genus Soft-shelled Turtle of the order Turtleidae of the order Reptiles, and is commonly known as soft-shelled turtle, turtle, and tortoise. It is distributed in various provinces in my country except Xinjiang, Tibet, and Qinghai. At present, the Chinese soft-shelled turtle breeding industry has developed into a large-scale and influential special aquaculture industry. In 2017, the total national ...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

Application Information

Patent Timeline
no application Login to view more
IPC IPC(8): C12Q1/6888C12Q1/6879C12Q1/686C12N15/11
Inventor 许晓军张海琪朱凝瑜孟庆辉
Owner ZHEJIANG FISHERIES TECH EXTENSION STATION
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Try Eureka
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap