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A nucleic acid library construction method and its application in the analysis of abnormal chromosomal structure of preimplantation embryos

A construction method and nucleic acid library technology, which is applied in the construction of nucleic acid library and the analysis of abnormal chromosome structure of pre-implantation embryos, and can solve the problems of long detection cycle, unfavorable clinical application promotion, and analysis errors, etc.

Active Publication Date: 2021-01-15
SUZHOU BASECARE MEDICAL DEVICE CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

This traditional method has a long detection cycle and a high risk of failure, which is not conducive to the promotion of clinical applications. At the same time, because it only captures and sequences a small number of SNP sites upstream and downstream of genetic disease genes, it is very easy to cause analysis errors due to homologous recombination.

Method used

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  • A nucleic acid library construction method and its application in the analysis of abnormal chromosomal structure of preimplantation embryos
  • A nucleic acid library construction method and its application in the analysis of abnormal chromosomal structure of preimplantation embryos
  • A nucleic acid library construction method and its application in the analysis of abnormal chromosomal structure of preimplantation embryos

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0071] Example 1 Balanced Translocation Genetic Family Embryo Detection

[0072] A family of chromosomal balanced translocation carriers who underwent assisted reproduction was recruited (the chip test results were available), and the family information is shown in Table 3. 5 mL of peripheral blood samples (a total of three blood samples) were collected from the balanced translocation carrier, wife, and carrier inherited from one parent, and stored in EDTA anticoagulated blood collection tubes. augmented product. Genomic DNA of the pedigree was extracted using a whole blood extraction kit. Preimplantation chromosomal balanced translocation analysis was performed using the method of the present invention.

[0073] Table 3 Chromosomal karyotype table of balanced translocation family (chip results)

[0074]

[0075] principle such as figure 1 As shown, firstly, a group of endonucleases recognize specific enzyme cutting sites on the genome, and the genomic DNA is cut into t...

Embodiment 2

[0126] Chromosomal aneuploidy cell line samples (5 cells) are used for details, see Table 12 below, and whole genome amplification is performed using the MDA method.

[0127] Table 12 Cell line sample details

[0128]

[0129] Detect according to the method of embodiment 1, cell line PGT-A data analysis result is as follows Figure 10 As shown, the detection results show that the method can detect more than 5M deletions.

Embodiment 3

[0136] The method of this embodiment is basically the same as that of Example 1, the only difference is that the endonuclease combination adopts BfaI and TaqI, or MboI and MspI respectively, and can also obtain sufficient SNP sites and indel sites under low data volume. Construct a genetic map to identify whether there is an abnormality in the structure of the embryo's chromosomes, and then distinguish between balanced translocation carrier embryos and normal embryos.

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Abstract

The invention relates to a method for constructing a nucleic acid library and its application in the analysis of abnormal chromosome structure of pre-implantation embryos. The combination of the first endonuclease and the second endonuclease performs enzyme cutting and breaking, and captures a DNA sequence within a fixed fragment range. , and then sequence the captured specific sequence. When the average sequencing depth of the genome is more than 3×, SNP analysis can be performed on the whole genome, and embryo balance translocation can be detected through linkage analysis of family samples. This method reduces the amount of data required for whole genome sequencing, while ensuring effective SNP sites and their depth, increasing the number of SNP sites that can be constructed from haplotypes, and obtaining sufficient coverage of the entire genome with a very low amount of sequencing data. Genomes can be used to analyze haplotype SNPs and indels, and there is no need to design multiple PCR primers for SNPs, which greatly reduces the amount of data required and detection costs.

Description

technical field [0001] The invention relates to the field of molecular biology, in particular to a method for constructing a nucleic acid library and its application in analyzing abnormal chromosome structure of pre-implantation embryos. Background technique [0002] Simplified genome sequencing is a method that cuts genomic DNA with restriction endonucleases, uses high-throughput sequencing for specified parts, and obtains a large number of genetic polymorphism tag sequences, so as to fully display the sequencing strategy of the entire genome of a species. This method can reduce the complexity of the genome, the implementation process is simple, and the cost is saved. At the same time, the genetic polymorphism tags in the whole genome can be obtained without relying on the reference genome. Therefore, it is widely used in molecular marker development, genetic map construction, and gene / QTL mapping. And genome-wide association analysis, population genetic analysis and molecu...

Claims

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Application Information

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IPC IPC(8): C12Q1/6806C12Q1/6869C12Q1/6883C40B50/06
CPCC12Q1/6806C12Q1/6869C12Q1/6883C40B50/06C12Q2600/156C12Q2535/122C12Q2525/191C12N15/1093C12Q2521/301C12Q2531/113C12Q2563/179C12Q2525/155C12Q2533/107C12Q1/6855C12Q1/686
Inventor 赵丁丁冒燕孔令印梁波
Owner SUZHOU BASECARE MEDICAL DEVICE CO LTD
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