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Kit for detecting genotype of human chromosome 21 STR (short tandem repeat)

A technology for detecting reagents and chromosomes, applied in the field of molecular biology, can solve the problems that QF-PCR technology has not yet been applied, and achieve high diagnostic efficiency, high detection sensitivity, and strong specificity

Active Publication Date: 2013-05-15
ATTACHED OBSTETRICS & GYNECOLOGY OSPITAL MEDICALCOLLEGE ZHEJIANG UNIV +2
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although STR typing technology has been widely recognized in forensic DNA typing and other fields, it has not been applied in QF-PCR technology for detecting chromosome number

Method used

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  • Kit for detecting genotype of human chromosome 21 STR (short tandem repeat)
  • Kit for detecting genotype of human chromosome 21 STR (short tandem repeat)
  • Kit for detecting genotype of human chromosome 21 STR (short tandem repeat)

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0033] Embodiment 1: Application of the kit for detecting the STR genotype of human chromosome 21 in the detection of the number of chromosome 21

[0034] (1) Kit composition (100 people) (Table 3).

[0035]

[0036] (2) Main instruments and equipment

[0037] PCR amplification instrument, ABI310 genetic analyzer, high-speed centrifuge, biological safety cabinet or clean bench, micro pipette, ultraviolet spectrophotometer, constant temperature water bath, refrigerator, etc.

[0038] (3) DNA extraction

[0039] The sample is 0.2ml of fresh EDTA anticoagulated whole blood or 10ml of amniotic fluid, DNA is extracted by Chelex-100 method (Chelex reagent is Bia-rad product), the purity and concentration of DNA are detected by UV spectrophotometer, and the extracted sample DNA is added to pure water Dilute to a concentration of about 0.1ng / μL-0.5 ng / μL.

[0040] (4) PCR reaction system (25 μL) (Table 4).

[0041]

[0042] (5) PCR cycle parameters: 95°C for 2 minutes - (9...

Embodiment 2

[0049] Example 2: Rapid prenatal diagnosis of trisomy 21 with this kit

[0050] 558 remaining amniotic fluid samples (amniotic fluid volume 0.5-1ml) analyzed by conventional karyotype were used in the kit of the present invention for blind detection and analysis according to Example 1. A total of 5 cases of trisomy 21 (all complete types, no chimeras and translocations) were detected in 558 amniotic fluid samples by this kit, and the test results can be obtained within 48 hours for each sample.

[0051] Compared with the results of chromosome karyotype diagnosis, all trisomy 21 was detected by D21 trisomy analysis fluorescent detection kit, without missing detection and amplification failure, with 100% diagnostic sensitivity and 100% diagnostic accuracy. From the perspective of the detection time of a single sample, the time-consuming use of the D21 trisomy analysis fluorescent detection kit is only 1 / 10 of the traditional karyotype analysis method. Due to the high-throughp...

Embodiment 3

[0053] Example 3: Determining the source of extra chromosomes in trisomy 21 embryos

[0054] Collect 6 aborted villi tissues (about 2 g each) known to be trisomy 21 karyotype (already done karyotype analysis), and extract peripheral blood samples from the father and mother of the aborted embryo (2ml EDTA per case) Anticoagulant), use the kit of the present invention to carry out blind detection and analysis according to the implementation case 1. The results show that this kit can clearly determine the relative source of redundant chromosomes through detection, which is helpful for guiding the next pregnancy. attached Figure 5-a~5-c It is the analysis of the source of redundant chromosomes in trisomy 21 aborted embryos. It is the genotype detection of one family at the D21S11 and D21S1246 loci. Since the alleles of the embryos come from the parents, the genotype comparison shows that the embryo samples are at D21S11 , The redundant allele on the D21S1412 locus comes f...

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Abstract

The invention provides a kit for detecting genotype of human chromosome 21 STR (short tandem repeat). The kit comprises an amplification reagent and an amplification product detection reagent and mainly contains primers of 12 pairs of fluorescent markers, a molecular weight internal label containing the size of 14 fragments, an allele typing standard product and a quality control product. According to the kit disclosed by the invention, when the kit is used, the required quantity of specimens is small, the diagnosis is fast, and the kit is suitable for villi, amniotic fluid, blood and other tissues, and can be used for not only prenatal diagnosis, but also source analysis of extra chromosomes of a 21-trisomy aborted fetus; the kit can precisely diagnose the genotype and is conductive to presentation of diagnosis result and laboratory quality evaluation; and the kit can further realize fast, accurate, automatic and high-throughput detection of the number of chromosomes 21 and realize quality controllability and standardization of a detection result, and thus has clinical application prospects.

Description

technical field [0001] The invention belongs to the field of molecular biology, relates to fluorescent quantitative PCR technology (QF-PCR), in particular to a kit for detecting the STR genotype of human chromosome 21, which is a rapid and high-throughput detection of multiple STR locus genotypes. Background technique [0002] Trisomy 21 (also known as Down syndrome) is the most common chromosomal abnormality among live births, accounting for about 1 / 600. The number of chromosome 21 in patients with this disease has changed from two normally to three, and the clinical manifestations are severe mental retardation, prone to heart disease, leukemia, and low immune function. The disease currently has no treatment and is mainly prevented through prenatal diagnosis. The main method and gold standard of prenatal diagnosis carried out on a large scale in my country is cytogenetic karyotype analysis. This method is accurate and reliable, but it is based on manual operation, relying ...

Claims

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Application Information

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IPC IPC(8): C12Q1/68
Inventor 朱宇宁吕时铭薛佳郑卫国陈雁张民
Owner ATTACHED OBSTETRICS & GYNECOLOGY OSPITAL MEDICALCOLLEGE ZHEJIANG UNIV
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