Human x-chromosome dna 19 locus multiplex amplification kit and its application
An X chromosome and multiple amplification technology, which is applied in the field of multiple amplification kits for 19 loci of human X chromosome DNA, can solve problems such as complex calculations and inability to guarantee non-linkage, achieve high sensitivity, save time for testing personnel, and reduce the number of reagents. The effect of time on the ratio of PCR components
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Embodiment 1
[0035] Example 1 Determination of loci, primer design and establishment of kit amplification parameters
[0036] The selected loci mainly consider the following factors: 1. Polymorphism, the GD value of the selected STR loci is relatively high in the Chinese population > 0.6, 2. Select closely linked loci in different linkage groups, located in the same linkage group The loci are inherited in linkage and can be used to observe polymorphisms by haplotype. 3. Select loci with a relatively low mutation rate, such as DXS8377 and DXS10011 with a mutation rate of 3.7% and 2.5%, which cannot be selected. 4. Consider primer design factors, such as the DXS10148 template sequence, as follows, there are 14 polyA structures not far before the core repeat region, so the primer cannot span this region. Primers can only be in the back region, such as AAAAGGGGGAAGGAAGGAAG. The primer contains 3 repeats of GGAA, which is enough to match the locus of TTCC in the core repeat region to produce ...
Embodiment 2
[0062] Example 2 Identification of Sisterhood Using the Fluorescence-labeled Multiplexed Amplification Test System of 19 Loci
[0063] Such as image 3 , linked X loci have unique advantages in complex paternity testing, such as sisterhood determination. B and A may be half-sisters, whose biological father M has died, M1 and M2 are M's brothers, and M3 is M's sister.
[0064] 1. Collect blood samples in identification cases: The samples in this experiment are provided by XX paternity identification agency. For DNA extraction, use the Chelex-100 method to extract genomic DNA from two whole blood samples: take 0.5-5 μl of whole blood into a sterilized 1.5ml centrifuge tube, add sdH 2 O1ml in the tube, shake for a few seconds; place at room temperature for 10 minutes, shake for a few seconds, centrifuge at 12,000rpm for 3 minutes, discard the supernatant, keep enough supernatant to cover the precipitate, do not stir the precipitate; add 200μl of 5% Chelex-100 , shake for a few...
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