Unlock instant, AI-driven research and patent intelligence for your innovation.

Primer, probe and molecular detection method for quality control of nucleic acid sample of mammal

A mammalian, sample quality technology, applied in the determination/inspection of microorganisms, biochemical equipment and methods, DNA/RNA fragments, etc., can solve problems such as inability to monitor nucleic acid degradation and contamination

Active Publication Date: 2015-02-18
YANGZHOU UNIV
View PDF0 Cites 0 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the means of adding exogenous control genes cannot monitor the problems of nucleic acid degradation and contamination in sample collection, transportation, storage and other links
At present, there is no system in the world that monitors the nucleic acid level and quality of the samples to be tested and is conveniently used for PCR quality control.

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
  • Primer, probe and molecular detection method for quality control of nucleic acid sample of mammal
  • Primer, probe and molecular detection method for quality control of nucleic acid sample of mammal
  • Primer, probe and molecular detection method for quality control of nucleic acid sample of mammal

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0043] Example 1: Detection of HMBS copy number in 383 blood samples from 13 species in 9 countries.

[0044] Use the HMBS FRET-PCR system established by the present invention to detect the HMBS copy number of nucleic acids from 383 blood samples from 8 countries, people and 12 mammals, and experience different transportation and storage environments, which is a reasonable collection and preparation of samples. Storage and transportation provide a theoretical basis. The source, type and HMBS copy number of the samples are shown in Table 1. Of the whole blood collected from 50 dairy cows in China, HMBS gene was not detected in 18 cows, and the HMBS copy number of this group (5.8×10 4 ±2.3×10 5 ) was significantly lowest among the 383 samples tested. In addition, the HMBS copy number (1×10 5 ±9×10 4 ) were also significantly lower than similar blood samples collected in the Caribbean (Costa Rica: 2.3x10 6 ±2.5x10 6 ; Nicaragua: 1.7x10 6 ±8.8x10 5 ). A comprehensive com...

Embodiment 2

[0049] Example 2: Detection of HMBS copy numbers in 11 tissues and organs of mice.

[0050] Blood and 11 tissues and organs were collected from 8 BALB / c and C57BL / 6 mice (two positive and two negative mice for each breed) for nucleic acid purification and HMBS-PCR. The copy number of HMBS in mouse samples has small differences among mouse strains, sexes and various tissues and organs. HMBS copy number of female mice (10 2.67 ±10 0.53 ) significantly higher than that of male mice (10 2.37 ±10 0.68 ). HMBS copy number in the heart (10 2.98 ±10 0.26 ), liver (10 2.70 ±10 0.39 ), spleen (10 3.15 ±10 0.39 ), lungs (10 2.82 ±10 0.39 ) and kidneys (10 3.11 ±10 0.32 ) significantly higher than in fat (10 2.05 ±10 0.29 ), bones (10 1.49 ±10 0.67 ) and hair (10 1.73 ±10 0.24 ) HMBS copy number ( figure 2 ).

Embodiment 3

[0051] Example 3: Comparison of HMBS copy number of the present invention and PicoGreen method to determine the absolute content of nucleic acid.

[0052] The PicoGreen fluorescence method is widely used in the quantitative determination of the absolute content of double-stranded DNA. We compared the correlation of HMBS copy number and absolute DNA content of the present invention. The HMBS copy number and absolute DNA content were determined in blood samples collected from 39 dogs in Costa Rica and 40 dogs in Inner Mongolia, China. Blood samples collected in Costa Rica did not show hemolysis, and no freezing and thawing of samples occurred before nucleic acid purification. However, the blood samples collected in Inner Mongolia experienced several freezing and thawing, resulting in obvious hemolysis. Figure 4 As shown, the HMBS copy number of the Costa Rican sample that had not undergone freeze-thaw was significantly higher than that of the Inner Mongolia blood sample that ...

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 relates to a primer, a probe and a molecular detection method for quality control of a nucleic acid sample of a mammal. The primer and the probe are as shown in SEQ ID NO.1-4. The molecular detection method comprises the following steps: carrying out PCR (Polymerase Chain Reaction) amplification on a nucleic acid sample by using the primer and the probe, and quantifying and / or classifying the nucleic acid sample according to the change of fluorescence intensity in the PCR process and a strip of a PCR amplification product in agarose gel electrophoresis. The primer, the probe and the molecular detection method disclosed by the invention can be used for conveniently and effectively monitoring the potential problems appearing in any links, such as sampling collecting, transporting, storing and extracting and provide a technical support for improving the PCR credibility, reducing the false negative report rate and the like.

Description

technical field [0001] The invention establishes a set of endogenous gene system based on the hydroxymethylcholane synthetase gene and suitable for PCR quality control of mammals used. The system can highly sensitively and specifically amplify human, human and Nucleic acids from 12 mammals (monkey, cat, dog, cow, donkey, goat, sheep, horse, leopard, lion, pig and tiger), 11 tissues and organs (heart, liver, brain, spleen) from two mouse strains , lung, fat, kidney, muscle, hair, bone, skin) nucleic acid, but does not amplify the nucleic acid of bacteria, parasites, viruses, edible vegetable oil. In addition, the amplified fragment designed by the present invention contains a gene sequence region, which is significantly different from all mammals, and the sequencing of this region can quickly and conveniently identify the species of mammals. Background technique [0002] Since its invention in 1986, PCR technology has been widely used in various fields such as medicine, bio...

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
Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/68C12N15/11
CPCC12Q1/6851C12Q1/686C12Q2565/125C12Q2545/114
Inventor 王成明危蓝菁张继垒杨奕张真稳郁多男陶建平杨章平郝永清徐达陆光武庄林林
Owner YANGZHOU UNIV