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Method, device and use thereof for determining the proportion of free nucleic acid in a biological sample

A technology for free nucleic acid and biological samples, which is applied in the field of determining the proportion of free nucleic acid in biological samples, and can solve problems that need to be improved

Active Publication Date: 2019-08-09
BGI GENOMICS CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] However, current methods for determining the proportion of cell-free nucleic acids in biological samples still need to be improved

Method used

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  • Method, device and use thereof for determining the proportion of free nucleic acid in a biological sample
  • Method, device and use thereof for determining the proportion of free nucleic acid in a biological sample
  • Method, device and use thereof for determining the proportion of free nucleic acid in a biological sample

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0349] According to the method for determining the proportion of free nucleic acid in biological samples of the present invention, the proportion of free fetal DNA was estimated for 11 cases of plasma samples from pregnant women to be tested, as follows:

[0350] 1) Sample collection and processing

[0351] 2 ml of peripheral blood during pregnancy were extracted from 11 pregnant women to be tested and 37 pregnant women known to be pregnant with male fetuses for plasma separation, so as to obtain peripheral blood samples of each pregnant woman to be tested and peripheral blood samples of pregnant women known to be pregnant with male fetuses.

[0352] 2) Library construction

[0353] The library was constructed according to the requirements of Complete Genomics Inc. for plasma library construction.

[0354] 3) Sequencing

[0355] The sequencing process was carried out on the machine in strict accordance with the standard operating procedures of Complete Genomics Inc.

[0356...

Embodiment 2

[0368] According to the method of the present invention, based on the results of the free fetal DNA ratio determined in Example 1, the plasma of 11 pregnant women with twins described in Example 1 was detected for chromosomal aneuploidy:

[0369] Based on the results of the free source free fetal DNA ratio determined in Example 1, according to the following steps, determine the fetal chromosomal aneuploidy of 11 samples to be tested by fetal concentration:

[0370] a) Calculate the fetal concentration fra.chri through the i (i=13, 18, 21) chromosome;

[0371] b) Estimate the fetal concentration fra.size by fragment;

[0372] c) Judgment criteria:

[0373] I. When the value of fra.chri / fra.size is less than 0.35, the i-th chromosome of both fetuses in twins is normal;

[0374] II. When the value of fra.chri / fra.size is not less than 0.35 and not more than 0.7, one fetus in the twins has the i-th chromosome as a trisomy, and one fetus has normal i-th chromosome;

[0375] III....

Embodiment 3

[0400] Example 3: Chimera detection

[0401] In the following examples, the fragmented DNA of aborted tissue is mixed with the plasma of a non-pregnant woman according to a certain ratio as a simulated pregnant woman sample. Abnormal chromosome numbers (including complete trisomy, complete monosomy, mosaic in trisomy and mosaic in monosomy) are detected according to the following method, which includes the following steps:

[0402] 1) Whole Genome Sequencing (WGS): Use a high-throughput platform to perform whole genome sequencing on the samples to be tested.

[0403] 2) Obtain position information of Effective Reads: compare the sequencing sequence of the test sample with the reference genome sequence, and obtain the position information that can be uniquely compared to the reference sequence.

[0404] 3) Count the percentage of the unique alignment sequence of each chromosome and the percentage of G and C bases in the total bases of the unique alignment sequence of each chro...

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Abstract

The invention discloses a method, device and application for determining the proportion of free nucleic acid from a predetermined source in a biological sample, wherein the method includes: (1) performing nucleic acid sequencing on a biological sample containing free nucleic acid, so as to obtain multiple sequencing data result; (2) based on the sequencing results, determining the number of nucleic acid molecules in the sample whose length falls within a predetermined range; and (3) determining the number of nucleic acid molecules in the biological sample based on the number of nucleic acid molecules whose length falls within a predetermined range The proportion of free nucleic acid described in. The method can accurately and efficiently determine the proportion of free nucleic acid from a predetermined source in biological samples, and is especially suitable for determining the proportion of fetal nucleic acid in the peripheral blood of pregnant women and the proportion of tumor nucleic acid in the peripheral blood of tumor patients.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a method, device and application for determining the proportion of free nucleic acid in a biological sample. Background technique [0002] Since 1977, researchers have successively discovered cancer-derived DNA in the peripheral blood of tumor patients, and confirmed the existence of cff-DNA in the plasma of pregnant women, and detected and estimated cancer-derived DNA in the peripheral blood of tumor patients, as well as pregnant women. The fraction of cell-free fetal DNA in plasma, ie, the determination of the fraction of cell-free nucleic acids of predetermined origin in a biological sample, is significant. [0003] However, current methods for determining the proportion of cell-free nucleic acids in biological samples still need to be improved. Contents of the invention [0004] The present invention aims to solve at least one of the technical problems existing in the prior ar...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G16B10/00C12Q1/6869C12M1/00
CPCC12Q1/6869G16B20/20G16B20/10G16B30/10C12Q2535/119C12Q1/68G16B15/00G16B20/00G16B30/00G16B15/10
Inventor 蒋馥蔓袁玉英王威尹烨
Owner BGI GENOMICS CO LTD