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Primer set and detection kit for detecting STR gene locus of human chromosome 4 and application of primer set

A technology for chromosome and human detection, applied in recombinant DNA technology, biochemical equipment and methods, and microbial measurement/inspection, etc., can solve the problems of few STR detection sites and single quantity, and achieve fast detection speed, high sensitivity, The effect of improving detection efficiency

Active Publication Date: 2018-11-20
GUANGZHOU DARUI BIOTECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] At present, most of the common STR detection kits at home and abroad are developed for forensic identification and paternity identification. The selected STR loci are basically on different chromosomes, and most of the STR loci selected for a single chromosome are no more than 4 (some for X or Y chromosome detection kit), there are fewer detection kits covering the STR locus of chromosome 4, and fewer STR detection loci of chromosome 4 are included in the detection kit (mostly only 1 STR locus) However, STR detection kits for chromosome 4 alone have not been found. For example, QIGEN's Investigator HDplex Kit and Biotype's climera only detect the D4S2366 site in chromosome 4. The domestic Beijing Basic Cognitive Technology Co., Ltd. Also only D4S2366
Moreover, the STR loci of foreign kits do not necessarily have the polymorphism and specificity of the Chinese population. The domestic STR detection kits, the STR loci of chromosome 4 are similar to those of foreign countries and the number is single

Method used

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  • Primer set and detection kit for detecting STR gene locus of human chromosome 4 and application of primer set
  • Primer set and detection kit for detecting STR gene locus of human chromosome 4 and application of primer set
  • Primer set and detection kit for detecting STR gene locus of human chromosome 4 and application of primer set

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0070] Example 1: Optimization of STR loci and their primers that preferably conform to the polymorphisms of the Han population

[0071] The kit of the present invention is developed for the Chinese population, and the STR sites with polymorphisms in the Chinese Han population are screened through the following specific implementation steps, and primers are designed and optimized for their sequences:

[0072] 1. Preliminary screening of STR loci

[0073] Select 20 STR loci of chromosome 4 (D4S2430, D4S174, D4S3268, D4S421, D4S2381, D4S3332, D4S2620, D4S1563, D4S2622, D4S3248, D4S2366, D3362, D4S3359, 2 , D4S98, D4S96, D4S3351, D4S3329, D4S2394), search for the nucleotide sequence of the STR locus in the NCBI library, use SSRHunter software for sequence analysis, and obtain candidate STR loci by screening according to the following principles: (1) STR is easy Good amplification, analysis, and stability (2) The STR loci of the same chromosome are distributed as much as possible...

Embodiment 2

[0078] Embodiment 2: the use of detection kit

[0079] Prepare a kit including the following components: PCR components, PCR mixture MIX, 1 tube (1100 μL / tube); 10× primer mixture, 1 tube (30 μL / tube); PCR enzyme system, 1 tube (75 μL / tube); tube); positive quality control product for trisomy 4 determined by karyotype analysis, 1 tube (20 μL / tube); STR negative quality control product confirmed by karyotype analysis with normal ploidy, 1 tube (20 μL / tube).

[0080] 1. Specimen collection, transportation and storage

[0081] (1) Specimen collection: Specimens can be blood, amniotic fluid, villi tissue, aborted embryo tissue, etc. 2mL of venous blood or 0.5-1mL of fetal umbilical cord blood was taken routinely, and anticoagulated with EDTA; 2-5mL of amniotic fluid or several strips of villous tissue (100-500mg) were obtained by puncture.

[0082] (2) Storage: Specimens can be tested immediately, stored at 4°C for one week, and stored at -20±5°C for up to one year.

[0083] (3...

Embodiment 3

[0103] Example 3: Detection of spontaneous abortion samples (4-trisomy) by using this kit to amplify

[0104] Abortion tissue samples were used for blind detection and analysis according to the method of implementation case 1. The results showed that the test kit obtained the test results within 48 hours, and 3 cases of trisomy of chromosome 4 were detected (case 1, case 2 and case 3). Its sensitivity and diagnostic accuracy are 100%. QF-PCR is combined with capillary electrophoresis technology to achieve high-throughput detection and higher efficiency. For specific results, see the attached image 3 , with Figure 4 , with Figure 5 Shown: Example 1 (attached image 3 ) in the loci of the three peaks have D4S3351, D4S3248 and D4S2381 loci, and the peak areas of the other loci are also approximately 1:2 or 2:1; Example 2 (attached Figure 4 ) in the loci of the three peaks have D4S3248 and D4S2381 loci, and the peak areas of the other loci are also approximately 1:2 or 2:1...

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Abstract

The invention discloses a primer set and a kit for detecting an STR genotype of a human chromosome 4 and application of the primer set. The primer set comprises five primers of an STR gene locus of achromosome 4, specifically as follows: D4S2381, D4S2620, D4S3248, D4S2622 and D4S3351. The kit provided by the invention develops a detection kit for analyzing chromosomal ploidy specific to the STR gene locus of the chromosome 4 with polymorphism and specificity of Chinese population on account of a current status of lacking the detection kit special for analyzing the STR gene locus of the chromosome 4; and the detection kit provided by the invention is wide in application range, suitable for analyzing spontaneous abortion samples caused by abnormal number of the chromosomes 4, and also ableto analyze abnormality of the chromosome 4 in prenatal diagnosis.

Description

technical field [0001] The invention relates to the field of gene detection, which is a kit for detecting the STR locus type of chromosome 4 in human autosomes by using multiple fluorescent PCR technology combined with capillary electrophoresis technology. Background technique [0002] Human trisomy 4 is more common in acute nonlymphocytic leukemia and lymphoproliferative disorders, and has also been found in spontaneously aborted fetuses. In individual studies, chromosome 4 was found to be associated with Parkinson's disease and visual abnormalities. In addition, trisomy of chromosome 4 may be related to placental development. Types of chromosomal abnormalities include abnormal chromosome number and abnormal chromosome structure. Spontaneous abortion accounts for about 10% to 15% of clinical pregnancies. Among spontaneous abortions caused by chromosomal abnormalities, about 96% are numerical abnormalities, and only 3% to 5% are structural abnormalities, among which there ...

Claims

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Application Information

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IPC IPC(8): C12Q1/6858C12N15/11
CPCC12Q1/6858C12Q2537/143C12Q2565/125C12Q2521/531C12Q2563/107C12Q2527/137
Inventor 吴英松周其伟李明杨学习康小龙何丹
Owner GUANGZHOU DARUI BIOTECH
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