Eight-color system STR (short tandem repeat) site multiplex amplification kit aiming at difficult detection material and application of eight-color system STR site multiplex amplification kit

A compound amplification and kit technology, applied in the field of molecular biology, can solve the problems such as the inability to meet the requirements of higher identification ability, the limited number of fluorescence channels of genetic analyzers, poor complementarity and versatility, etc., and achieve amplification of fragments. The effect of short, good polymorphism and short amplification time

Pending Publication Date: 2022-05-03
苏州市公安局 +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

ABI and Promega have launched two kits, MiniFiler and S5 System, respectively, but they both have great limitations, fewer sites, and poor complementarity and versatility in joint application with other commercial kits
Based on the six-color fluorescent system in China, Wuxi Zhongde Meilian Company also launched a Mini-STR product with 16 sites. Although the individual recognition efficiency has reached the same level as Identifiler, it is limited by the number of fluorescent channels of the genetic analyzer. The kit still cannot meet the customer's requirements for more sites and higher recognition ability

Method used

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  • Eight-color system STR (short tandem repeat) site multiplex amplification kit aiming at difficult detection material and application of eight-color system STR site multiplex amplification kit
  • Eight-color system STR (short tandem repeat) site multiplex amplification kit aiming at difficult detection material and application of eight-color system STR site multiplex amplification kit
  • Eight-color system STR (short tandem repeat) site multiplex amplification kit aiming at difficult detection material and application of eight-color system STR site multiplex amplification kit

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0033] 30pg and 0.5ng standard product 9948 were detected using the kit of the present invention.

[0034] The kit of the present invention consists of the following: primer mixture, 1mL; PCR buffer Unique Mix, 2mL; sdH 2 O, 1.95 mL; PCR enhancer.

[0035] The composition of the PCR buffer Unique Mix is: hot start Taq enzyme 0.2U / μL~0.5U / μL, Tris-HCl 50mM~125mM with pH 8.25~8.4, dNTPs 5.0mM~7.5mM, KCl 50mM~125mM, BSA 2mg / ml~5mg / ml.

[0036] The optional types of PCR enhancers are: polyethylene glycol, Triton X-100, polyvinylpyrrolidone (PVP) or dithiothreitol, and one or two of them are selected and directly added to the PCR buffer Unique Mix.

[0037] 1. The operation steps are as follows:

[0038] 1.1 Amplification system

[0039] Component volume Unique Mix 10.0 μL 30pg / 0.5ng9948 1μL Primer set 5.0 μL wxya 2 o

Make up to 25 μL

[0040] 1.2 The amplification procedure is:

[0041]

[0042] 1.3. Fluorescent detection of...

Embodiment 2

[0047] A total of 24 samples of samples such as saliva spots, mouse swab samples, and old degraded samples in actual cases were detected using the kit of the present invention.

[0048] The kit of the present invention consists of the following: primer mixture, 1mL; PCR buffer Unique Mix, 2mL; sdH 2 O, 1.95 mL; PCR enhancer.

[0049] The composition of the PCR buffer Unique Mix is: hot start Taq enzyme 0.2U / μL~0.5U / μL, Tris-HCl 50mM~125mM with pH 8.25~8.4, dNTPs 5.0mM~7.5mM, KCl 50mM~125mM, BSA 2mg / ml~5mg / ml.

[0050] The optional types of PCR enhancers are: polyethylene glycol, Triton X-100, polyvinylpyrrolidone (PVP) or dithiothreitol, and one or two of them are selected and directly added to the PCR buffer Unique Mix.

[0051] 1. Sample preparation: extract by magnetic bead method, refer to "GA / T 383-2002 Forensic Science DNA Laboratory Inspection Specification" for specific methods.

[0052] 2. The operation steps are as follows:

[0053] 2.1 Amplification system

[0...

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Abstract

The invention discloses an eight-color system STR (short tandem repeat) locus multiplex amplification kit for a difficult sample and application, the kit comprises specific amplification primers of 26 STR loci, and the STR loci comprise CSF1PO (cytoplasmic globulin A), FGA (fibroblast growth factor A), TH01, TPOX, vWA, D3S1358, D5S818, D7S820, D8S1179, D13S317, D16S539, D18S51, D21S11, Penta D, Penta E, D6S1043, Amelogenin, D2S1338, D12S391, D19S433, D1S1656, D2S441, D22S1045 and D10S1248, as well as two sex The maximum amplification enhancers of all the sites are within 300bp. The kit provided by the invention has the advantage of rapid and efficient detection of difficult detection materials such as low copy, high degradation and high inhibitors in on-site cases; all gene loci are compatible with an existing case database; and the time of case checking is shortened, the public security officers are helped to quickly lock the suspects, and the case cracking efficiency is improved.

Description

technical field [0001] The invention belongs to the field of molecular biology, and relates to the use of chemical fluorescent molecular marker primers for gene detection, in particular to an eight-color system STR site composite amplification kit for difficult inspection materials and its application. Background technique [0002] At present, the fluorescent labeling STR typing kits commonly used in forensic practice are from Applied Biosystems (ABI) and Promega. ABI recently launched the VeriFilerPlus PCR amplification kit, which is a six-color fluorescent system, including 23 autosomal loci, 2 sex loci Amelogenin and Y-Indel, and the size range of the amplified product is 80bp-430bp. Promega's new case test is the VersaPlex 27PYSTR kit, which is also a six-color fluorescent system, containing 23 autosomal loci, 1 sex locus Amelogenin, and 3 Y-STR loci (DYS391, DYS576, and DYS570) , the size range of the amplified product is 75bp-480bp. When the above two STR kits detect...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6888C12Q1/686C12N15/11
CPCC12Q1/6888C12Q1/686C12Q2600/156C12Q2563/107
Inventor 周如华王鑫石云杰张健陈维忠崔扬邵泽香梅兴林李娟史明皓姜颖晔范晓芸
Owner 苏州市公安局
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