Looking for breakthrough ideas for innovation challenges? Try Patsnap Eureka!

Novel method and kit for extracting high-quality RNA (Ribose Nucleic Acid) from paraffin-embedded tissue

A paraffin-embedded, kit technology, applied in recombinant DNA technology, DNA preparation and other directions, can solve the problems of small RNA fragments, difficult RNA extraction, easy degradation, etc., achieve long RNA length, save research funds, species information full effect

Inactive Publication Date: 2011-07-06
GENERAL HOSPITAL OF TIANJIN MEDICAL UNIV
View PDF0 Cites 9 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] The purpose of the present invention is to provide a new method and kit that can simply and effectively extract high-quality RNA from conventional pathological paraffin tissues, so as to solve the problem of RNA extraction when studying conventional formalin-fixed and paraffin-embedded tissues by molecular means. It is difficult to improve, easy to degrade, small RNA fragments and other defects

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
  • Novel method and kit for extracting high-quality RNA (Ribose Nucleic Acid) from paraffin-embedded tissue
  • Novel method and kit for extracting high-quality RNA (Ribose Nucleic Acid) from paraffin-embedded tissue
  • Novel method and kit for extracting high-quality RNA (Ribose Nucleic Acid) from paraffin-embedded tissue

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0034] Example 1 Using the method of the present invention to extract RNA from multiple lung cancer tissues

[0035] The kit formula is the preferred formula:

[0036] ⑴Reagent A: The ingredient is a non-toxic, environmentally friendly and transparent dewaxing agent;

[0037] ⑵Reagent B: pH8.0 10mmol / L Tris-HCl, 2%SDS, 0.1 mmol / L EDTA;

[0038] (3) Reagent C: proteinase K at a concentration of 60 mg / ml;

[0039] (4) Reagent D: It is made by mixing anhydrous phenol and chloroform at a volume ratio of 7:3;

[0040] (5) Reagent E: pH 5.3, sodium acetate with a concentration of 3mol / L;

[0041] (6) Reagent F: Glycogen with a concentration of 20ug / mL.

[0042] The extraction method is:

[0043] (1) Take 1-5 paraffin sections and place them in a 2 mL Eppendorf tube, add 1.5 mL of reagent A and mix well, incubate in a 37 oC water bath for 20 min, centrifuge at 10000 rpm for 6-10 min, discard the supernatant; repeat the above steps once;

[0044] (2) Add 1.5mL 100% ethanol, incu...

Embodiment 2

[0051] Example 2 Using the RNA extracted by the method of the present invention to amplify target genes of different fragment sizes

[0052] The total RNA of paraffin tissue obtained by the method of the present invention is in a diffuse state due to different degrees of degradation. In order to compare the effect of this method on the amplification of genes with different fragment sizes, frozen tissues and paraffin-embedded tissues were selected respectively in this example. The -actin gene was used as the object of amplification, and 13 different fragments of the β-actin gene were amplified. The specific operation is as follows:

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 novel method and a kit for extracting high-quality RNA (Ribose Nucleic Acid) from a paraffin-embedded tissue. The method comprises the following steps of: firstly, removing paraffin from a sample by using a nontoxic and non-volatile paraffin removing agent to improve the sensitivity of the paraffin-embedded tissue to lysate; secondly, cracking a tissue sample with SDS (Sodium Dodecyl Sulfate)-containing lysate and protease K, wherein the lysate provides a pH environment suitable for cell cracking, so that proteins are further denaturalized and water solubility is enhanced; thirdly, fully denaturalize the proteins with phenol-chloroform mixed liquid, separating the RNA and DNA (Deoxyribose Nucleic Acid) from the proteins, and chromatographing the RNA and the DNA from the mixed liquid to a water layer; fourthly, precipitating the RNA under the coaction of sodium acetate and glucogen; and lastly, dissolving the RNA in RNase-free water. By adopting the kit and the extraction method, the aim of obtaining total RNA with high quality, a long segment and high yield from the paraffin-embedded tissue is fulfilled. The obtained RNA can be applied to medical molecular biology researches, clinical examination and the like.

Description

【Technical field】 [0001] The invention relates to a new method and reagent for extracting high-quality total RNA from paraffin-embedded conventional pathological tissues. 【Background technique】 [0002] The occurrence of diseases is often accompanied by changes in gene expression status. Comprehensive analysis of gene expression by extracting RNA from fresh or frozen tumor tissues is playing an increasingly important role in understanding disease occurrence, classification, and prognosis. However, there are limited sources of fresh or frozen tissue specimens. In contrast, formalin-fixed and paraffin-embedded tissues come from a wide range of sources, and the pathology departments of major hospitals have a large number of archives. On the other hand, this part of the specimens often has good follow-up data, which makes it possible to conduct large-scale clinical retrospective studies using archived wax blocks. [0003] However, compared with fresh tissue, RNA extraction in...

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): C12N15/10
Inventor 陈军周清华刘红雨李永文李颖
Owner GENERAL HOSPITAL OF TIANJIN MEDICAL UNIV
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Patsnap Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Patsnap Eureka Blog
Learn More
PatSnap group products

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

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap|About US| Contact US: help@patsnap.com