A next-generation sequencing-based method for STR typing of the locus D8S1179
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A technology of D8S1179STR and D8S1179, applied in the field of STR typing of the locus D8S1179, can solve the problems of allele loss, unbalanced amplification, limited genetic information, etc.
Active Publication Date: 2020-03-31
BEIJING CENT FOR PHYSICAL & CHEM ANALYSIS
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[0003] However, with the continuous improvement of DNA extraction ability and detection technology sensitivity, forensic physical evidence detection has developed from constant detection to trace or even trace detection. In the practice of physical evidence detection, the inherent shadow peaks of STR typing technology, Heterozygous locus amplification imbalance (heterozygote peak imbalance), allele loss (allelic drop-out), and low copy number template low copy number) make the genetic information obtained by traditional STR typing technology limited. There are application limitations in physical evidence inspection, and it is difficult to satisfactorily solve trace or complex problems in individual identification
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Embodiment 1
[0031] Example 1 Amplification of D8S1179 Gene Specific Fragment
[0032] With the consent of the blood donor, normal blood was collected with an anticoagulant tube, and the blood collection tube was immediately placed in an ice box. All blood samples were analyzed by molecular biology method, compared with the human genome sequence reported in NCBI database, PCR method was used, and primers were designed in the conserved sequence region of D8S1179 gene using Primer Express Version 3 software according to the principles of primer design:
[0035] 1. Use Chelex100 kit to extract DNA from blood of different samples:
[0036] Cut an appropriate amount of blood spots, put them in a centrifuge tube for 0.5 minutes, add an appropriate amount of pure water, soak at room temperature, centrifuge at 13,000 rpm for 5 minutes, discard the supernatant, leave about 20 μL of liquid ...
Embodiment 2
[0063] Example 2 Establishment of a method for analyzing biological information of the D8S1179 gene by next-generation sequencing
[0064] After the library is successfully constructed, take out all the sequencing sequences containing the sample bacode in the original data, first count the length distribution, distinguish the DNA sequence of each sample according to the index sequence in the primer of the sample, count the number of repetitions of the same DNA molecule, and find out the occurrence of SNPs, flanking sequences were retrieved and analyzed. The results shown in Table 2 can appear:
[0065] Table 2
[0066]
[0067] It can be seen that there are 12 and 13 repeats in this sample and in the second repeat, a SNP site with A mutation to G appeared.
[0068] Embodiment 3 Application experiment 1
[0069] For 9 samples of the existing first-generation typing results, extract DNA, use the primers described in the present invention, carry out the PCR method according...
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Abstract
The invention relates to the field of molecular biology and particularly discloses a specific primer pair for amplifying a gene D8S1179. The specific primer pair comprises D8S1179-FP:5'-TTTGTATTTCATGTGTACATTCGTAATTC-3' and D8S1179-RP:5'-ACCTATCCTGTAGATTATTTTCACTGATTG-3'. The invention further provides an STR typing method for a loca D8S1179 based on next generation sequencing and particularly provides a primer combination for being connected with a proper adaptor. The STR typing method is applied to the STR typing of the loca D8S1179. According to the primer pair capable of amplifying the gene D8S1179, after concentrations of different sample products subjected to PCR amplification, the equivalent mixing is carried out, and a genome library is constructed according to a BIOO RAPID DNA kit; the library is input into a computer by virtue of a next generation sequencer, data is subjected to bioinformatics analysis, and the typing results such as times of repetition of NDA molecules with same length, SNP of the loca and the difference among side wings of the loca of D8S1179 can be found.
Description
technical field [0001] The present invention relates to the field of molecular biology, in particular to a method for typing STRs based on the gene locus D8S1179 of next-generation sequencing. Background technique [0002] One of the basic tasks of forensic physical evidence is to solve the problem of individual identification. Forensic DNA analysis plays a vital role in clarifying the nature of the case and identifying the suspects, parties and victims. Forensic DNA analysis is a science and technology that uses modern DNA technology to analyze the distribution and transmission of DNA genetic markers in the population, to determine the consistency and genetic relationship of the analyzed samples, and to provide evidence for the investigation and judicial trial. Since 1985, Jeffereys and others established the first generation of typing technology - DNA fingerprinting technology, together with the later development of PCR amplified fragment length polymorphism analysis techn...
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