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Method for amplifying and detecting ribonucleic acid (RNA) fragments

A fragment and oligonucleotide technology, applied in DNA/RNA fragmentation, DNA preparation, recombinant DNA technology, etc., can solve the problems of rapid degradation, rare quantity, short fragment length, etc.

Pending Publication Date: 2022-03-04
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

These methods are limited due to the variety of implementations of the cfRNAs to be assessed, including rare numbers, short fragment lengths, high diversity, or rapid degradation

Method used

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  • Method for amplifying and detecting ribonucleic acid (RNA) fragments
  • Method for amplifying and detecting ribonucleic acid (RNA) fragments
  • Method for amplifying and detecting ribonucleic acid (RNA) fragments

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Embodiment

[0065] 1. Materials and methods

[0066] 1.1. Cell-free RNA isolation

[0067] Cell-free RNA was isolated from healthy male plasma. Whole blood samples from healthy males were collected in BD Vacutainer venous blood collection tubes (BD, #367525). Plasma fRNA fragments were isolated using the miRNeasy serum / plasma kit (Qiagen, #217184). Isolated cfRNA samples were quantified with Qubit RNA HS assay kit (Thermo Fisher, #Q32852) and stored at -70°C. Quantitative and qualitative evaluation of RNA and DNA samples was performed with Fragment Analyzer (AATI) using RNA or DNA gels.

[0068] 1.2 Conversion of cfRNA to cDNA and amplification to obtain dsDNA product

[0069] figure 1 A program showing the process of the present invention, comprising steps A to G.

[0070] Use the following steps to convert cfRNA samples to cDNA without purification.

[0071] Step A: 5' dephosphorylation of cfRNA

[0072] In Step A, the 5' end of the cfRNA is dephosphorylated. 5 μL of dephosphor...

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Abstract

The invention relates to a method for amplifying and detecting ribonucleic acid (RNA) fragments. In particular, the method of the present invention comprises converting an RNA fragment into cDNA and amplifying DNA. The invention also provides a kit for performing the method described herein.

Description

[0001] related application [0002] This application claims the benefit of U.S. Patent Provisional Application No. 62 / 850,651, filed May 21, 2019, based on 35 U.S.C. § 119, the entire contents of which are hereby incorporated by reference into this application. technical field [0003] The invention relates to a method for amplifying and detecting ribonucleic acid (RNA) fragments. Specifically, the method of the present invention comprises converting RNA fragments into cDNA and amplifying DNA. The invention also provides a kit for performing the methods described herein. Background technique [0004] RNA is an important genetic material involved in gene expression and regulation. Specifically, cell-free RNAs (cfRNAs) in biological fluids (e.g., blood, saliva, urine, etc.) have important genetic information of biological and medical significance, and thus, become valuable non-invasive samples for the diagnosis of many diseases. However, cfRNAs are very diverse and their st...

Claims

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Application Information

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IPC IPC(8): A61K31/713C07H21/02C07H21/04C12N15/10C12N15/11C12N15/113
CPCC12N15/1096C12Q1/6855C12Q2525/191C12Q2521/107C12Q2525/155C12Q2525/173C12N9/22C12Q1/686
Inventor 邱国平萧欣杰吴梓康
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