Method for detecting chromosome number variation of fetus

A technology of chromosome number and fetus, applied in the field of chromosome detection, can solve the problems of PCR amplification bias, uneven GC content of the genome, affecting the PCR amplification efficiency, etc., so as to reduce the investment of reagents, primers and time, and save the Effects of PCR amplification procedures, simplified library construction process

Active Publication Date: 2015-06-10
北京明谛生物医药科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The GC content of the human genome is not uniform, and factors such as complex secondary structures will affect the efficiency of PCR amplification, which will cause bias in PCR amplification and affect the accuracy of the detection structure

Method used

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  • Method for detecting chromosome number variation of fetus
  • Method for detecting chromosome number variation of fetus
  • Method for detecting chromosome number variation of fetus

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0062] Example 1 Method for constructing sequencing library

[0063] According to one aspect of the present invention, the present invention provides a method for constructing a sequencing library, such as figure 1 As shown, according to an embodiment of the present invention, the method includes the following steps:

[0064] Step S101, extract peripheral blood to obtain DNA in plasma. The term "DNA" used in the present invention can be any polymer comprising any deoxyribonucleotides, including but not limited to modified or unmodified DNA. Those skilled in the art can understand that the source of DNA is not particularly limited, and can be obtained from any possible way, it can be obtained directly from the market, it can also be obtained directly from other laboratories, and it can also be obtained directly from samples. extract from. According to an embodiment of the present invention, genomic DNA can be extracted from a sample. According to an embodiment of the presen...

Embodiment 2

[0069] Example 2 kit

[0070] According to another aspect of the present invention, the present invention provides a sequencing library kit for constructing a specific region of the genome of a sample. According to an embodiment of the present invention, the kit may include an enzyme for DNA end repair plus A base And the required buffer containing Tris-HCl, KCl, MgCl2, DDT, dNTPs; the ligase used to connect the modified DNA and the sequencing adapter and the required buffer containing Tris-HCl, MgCl2, DTT, ATP, betaine Buffers, connectors. Those skilled in the art can understand that the kit may further include any other components required for constructing the sample sequencing library.

[0071]The method for constructing a sequencing library of the present invention omits the step of PCR amplification, so the method for constructing a sequencing library of the present invention reduces the deviation generated by PCR amplification and improves the accuracy of sequencing. Be...

Embodiment 3

[0073] Example 3 The specific operation process of constructing the sequencing library

[0074] 1. Preparation of plasma DNA

[0075] The peripheral blood of 10 pregnant women was drawn, and the numbers were: 14N0001, 14N0002, 14N0003, 14N0004, 14N0005, 14N0006, 14N0007, 14N0008, 14N0009 and 14N0010. Blood samples were centrifuged in two steps to obtain plasma without blood cells, and sterilized high-purity water was used instead of plasma as a negative control, and the gold standard amniocentesis confirmed sample was used as a positive control, and the volume of each sample was 1ml.

[0076] Use the serum / plasma free DNA extraction kit produced by TIANGEN to extract DNA in plasma (QIAamp circulating nucleic acid kit from Qiagen or other companies’ DNA extraction kits are also applicable to the present invention), and the operation process is carried out according to the instructions.

[0077] 2. Repair of plasma DNA ends and addition of base A

[0078] The extracted DNA is ...

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Abstract

The invention discloses a reagent kit for detecting chromosome number of fetus and a method for detecting chromosome number of fetus that belong to the field of chromosome detection. The reagent kit comprises a reagent I that modifies DNA, phosphorylated sequencing linkers for linking two terminals of the modified DNA, a reagent II for linking the DNA phosphorylated sequencing linkers together, and a device for directly purifying the ligation product without amplifying by a primer. The provided method for detecting chromosome number of fetus comprises four steps, including a DNA extraction step, a DNA terminal modification step, a sequencing linker connection step, and a purification step. A sequencing library can be acquired without a PCR amplification. The disclosed reagent kit and the prepared library are able to achieve an efficient, precise and non-invasive effect for the detection of chromosome number of fetus.

Description

technical field [0001] The invention relates to the field of chromosome detection, in particular to a method for non-invasive detection of fetal chromosome number variation. Background technique [0002] Chromosomal aneuploidy is a disease caused by congenital abnormalities in the number of chromosomes, and it is one of the main diseases that cause birth defects in newborns. The abnormal number of chromosomes will lead to the increase or deletion of many genes, disrupt the balance of genes, and hinder the differentiation and development of related organs in the human body. The clinical manifestations are organ deformities, mental retardation, and growth retardation. Chromosomal aneuploidy diseases are divided into autosomal aneuploidy diseases and sex chromosome aneuploidy diseases. Trisomy 21 (Down Syndrome), Trisomy 18 (Edwards Syndrome), and Trisomy 13 (Patau Syndrome) are the three most common autosomal aneuploidies in neonates The incidence rates were 1 / 800, 1 / 8000 an...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68C12N15/10C40B50/06
Inventor 刘建云
Owner 北京明谛生物医药科技有限公司
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