Long distance PCR amplification method

A long-distance, reaction system technology, applied in the field of molecular biology, can solve problems affecting long-distance PCR amplification, and achieve the effect of less material requirements, low drug costs, and low equipment requirements

Inactive Publication Date: 2005-06-29
CHENGDU INST OF BIOLOGY CHINESE ACAD OF S
View PDF0 Cites 3 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Additionally, methodological issues and thermal cycling condition

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

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0041] Embodiment 1: Mitochondrial DNA (mtDNA) long-distance PCR amplification

[0042]Experimental material: Frozen whole material (stored at -20°C after freezing in liquid nitrogen): Scutigerruginosus, specimen number XM956, XM957, XM958, XM972. The specimens are preserved in the Amphibians and Herpetology Laboratory of Chengdu Institute of Biology, Chinese Academy of Sciences.

[0043] Muscle tissue mtDNA extraction: 10g of frozen muscle tissue, cut into pieces, put into a mortar, add liquid nitrogen and quickly grind into powder; add 50ml of SE solution, mix well, transfer to six 10ml centrifuge tubes; pipette, stand for 10min; Repeat pipetting 3 times; let stand for 10min; carefully absorb the supernatant, transfer to several 2ml Eppendorf tubes, centrifuge at 1000g for 10min, discard the precipitate, transfer the supernatant to a new Eppendorf tube, centrifuge at 12000g for 15min, discard the supernatant; collect Precipitate to a 2ml Eppendorf tube, add PBS buffer to wa...

Embodiment 2

[0092] Example 2: Amplification of lambda DNA.

[0093] Using λDNA as a template (Promega, Shanghai), primers, and reaction system are the same as in Example 1, except that the extension time of 30 thermal cycles is 38 min, and other thermal cycle parameters are the same as in Example 1. Electrophoresis detection showed that a 35kb DNA fragment was amplified.

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 belongs to the field of molecular biology, and discloses a long-distance PCR amplification method for difficulties such as harsh amplification conditions of longer gene sequences. The amplification method includes the following steps: designing primers, establishing a reaction system, and synthesizing After the primers are added to the established reaction system, the thermal cycle reaction is carried out under certain conditions. After the reaction is completed, the size of the amplified molecule is detected by electrophoresis to confirm that the amplification of the target fragment is completed. The present invention has the advantages of low cost, less materials, etc. advantage.

Description

1. Technical field [0001] The invention belongs to the DNA thermal cycle amplification method in the field of molecular biology. 2. Background technology [0002] Long-distance PCR amplification technology is the basis for further analysis of sequence structural genomics, which can amplify fragments of more than 20kb from the nuclear genome, including isolating complete genes from cDNA (Rose, 1991; Ohler and Rose, 1992; Ohler et al. al., 1993); including the analysis of high molecular weight restriction fragment length polymorphisms in a population (Ohler and Rose, 1992; Ohler et al., 1993). Many confusing questions in life science research will eventually be answered by the start of nuclear genome research. [0003] The advantage of long-distance PCR is that it requires less tissue material, and it can amplify a sufficient number of copies of the target gene DNA sequence from a sample of 1 ng or less (less than 300 copies, which can theoretically b...

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): C12P19/34
Inventor 莫邦辉曾晓茂郭骁才
Owner CHENGDU INST OF BIOLOGY CHINESE ACAD OF S
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