New Short Nucleotide Tandem Repeat Sequence Site and Its Application

Abstract

The present invention discloses a new short nucleotide tandem repeat locus and its application, the short tandem repeat locus G4S0001, the sequence is shown in SEQ ID NO.1, used to prepare (a) for genetic relationship analysis (b) Reagents or kits for individual identification; (c) Reagents or kits for paternity testing or blood relationship analysis; (d) Used to detect whether there is maternal blood contamination in the extracted amniotic fluid and / or (e) a kit for detecting whether leukocytes in a recipient are replaced by donor cells after bone marrow transplantation. The short tandem repeat locus G4S0001 has a high degree of discrimination and can effectively analyze genetic relationships.

Description

technical field

[0001] The invention relates to a short nucleotide tandem repeat sequence site and its application. Background technique

[0002] Short tandem repeat (Short Tandem Repeat.STR), also known as microsatellite DNA (microsatellite DNA), is a type of DNA tandem repeat sequence with a repeat unit of 2-6bp, a repeat number of 10-60, and a fragment length of less than 400bp; its cause Sliding during DNA replication, or base mismatch between the sliding strand and the complementary strand during replication and repair, resulting in the deletion or insertion of one or several repeat units. STR has a wide distribution in the genome, multiple alleles, high heterozygosity, and stable typing results. STR inheritance conforms to Mendelian inheritance laws (Koreth, J., et al. 1996. Microsatellites and PCR ge-nomic analysis. J. Pathol.178:239-248.) and other characteristics, and when multiple STR loci are jointly detected, the individual recognition ability and non-parent exc...

Examples

Embodiment 1

[0130] Combining the new STR locus G4S0001 of the present invention with 10 commonly used loci (VWA, D16S539, D5S818, D7S820, D13S317, D8S1179, D18S51, TH01, D2S1338, AMELO), multiplex fluorescent PCR technology was used to check the samples, and the four used The samples were three samples from a related family and one unrelated control sample. The primer sequences of G4S0001 and 10 commonly used loci are shown in Table 2.

[0131] Table 2 Primer Sequence

[0132]

[0133] Note: PET, VIC, NED, FAM four fluorescent dyes ((Applied Biosystems, USA company).

[0134] The specific experimental steps are as follows:

[0135] 1) DNA sample preparation

[0136] Collect blood from 4 selected sample individuals; extract DNA samples through DNA extraction kit and take 1 μl each, perform quality inspection and concentration estimation on the samples by 1% agarose electrophoresis, and then dilute the samples to the working concentration according to the estimated concentration 5-10...

Embodiment 2

[0148] Randomly select 20 groups of related and unrelated samples, and use the method in Example 1 for detection, the difference is that only the new STR locus G4S0001 of the present invention is used.

[0149] After testing, after determining the paternal sample, determine the offspring samples that are related to the male parent in the remaining 19 groups of samples. The new STR locus G4S0001 of the present invention can accurately exclude 12 groups of unrelated samples. A higher degree of discrimination was obtained.

technical field

[0001] The invention relates to a short nucleotide tandem repeat sequence site and its application. Background technique

[0002] Short tandem repeat (Short Tandem Repeat.STR), also known as microsatellite DNA (microsatellite DNA), is a type of DNA tandem repeat sequence with a repeat unit of 2-6bp, a repeat number of 10-60, and a fragment length of less than 400bp; its cause Sliding during DNA replication, or base mismatch between the sliding strand and the complementary strand during replication and repair, resulting in the deletion or insertion of one or several repeat units. STR has a wide distribution in the genome, multiple alleles, high heterozygosity, and stable typing results. STR inheritance conforms to Mendelian inheritance laws (Koreth, J., et al. 1996. Microsatellites and PCR ge-nomic analysis. J. Pathol.178:239-248.) and other characteristics, and when multiple STR loci are jointly detected, the individual recognition ability and non-parent exc...

Examples

Embodiment 1

[0130] Combining the new STR locus G4S0001 of the present invention with 10 commonly used loci (VWA, D16S539, D5S818, D7S820, D13S317, D8S1179, D18S51, TH01, D2S1338, AMELO), multiplex fluorescent PCR technology was used to check the samples, and the four used The samples were three samples from a related family and one unrelated control sample. The primer sequences of G4S0001 and 10 commonly used loci are shown in Table 2.

[0131] Table 2 Primer Sequence

[0132]

[0133] Note: PET, VIC, NED, FAM four fluorescent dyes ((Applied Biosystems, USA company).

[0134] The specific experimental steps are as follows:

[0135] 1) DNA sample preparation

[0136] Collect blood from 4 selected sample individuals; extract DNA samples through DNA extraction kit and take 1 μl each, perform quality inspection and concentration estimation on the samples by 1% agarose electrophoresis, and then dilute the samples to the working concentration according to the estimated concentration 5-10...

Embodiment 2

[0148] Randomly select 20 groups of related and unrelated samples, and use the method in Example 1 for detection, the difference is that only the new STR locus G4S0001 of the present invention is used.

[0149] After testing, after determining the paternal sample, determine the offspring samples that are related to the male parent in the remaining 19 groups of samples. The new STR locus G4S0001 of the present invention can accurately exclude 12 groups of unrelated samples. A higher degree of discrimination was obtained.

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