Kit, method and application for detecting mutation of predetermined locus in DNA sample

Kit, method and application for detecting mutation of predetermined locus in DNA sample

  • CN 102,409,089 B
  • Filed: 10/08/2011
  • Issued: 04/19/2017
  • Est. Priority Date: 10/08/2011
  • Status: Active Grant
First Claim
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1. it is a kind of for detecting DNA sample in deaf relevant mutational site combination system, it is characterised in that include:

  • DNA sample amplification device, the DNA sample amplification device, for entering performing PCR amplification to the DNA sample, to obtainAmplified production, the amplified production includes the mutational site;

    Amplified production extension apparatus, the amplified production extension apparatus is connected with the DNA sample amplification device, so as to from describedDNA sample amplification device receives amplified production, and carries out oligonucleotide extension to the amplified production, to obtainThe extension products of 3 '"'"' one base of end connection of the extension primer, wherein, 3 '"'"' ends of the extension primer are close to the mutationSite;

    Molecular weight detection device, the molecular weight detection device is connected with the amplified production extension apparatus, described to determineThe molecular weight of extension products;

    AndMutation analysises device, molecular weight of the mutation analysises device based on the extension products, determines the mutational siteMutation type,Wherein, detect that testing result when being the positive is with all mutational sites in the deaf relevant mutational site combinationStandard,Amplimer pair is provided with the DNA sample amplification device, extension is provided with the amplified production extension apparatus and is drawnThing,The mutation for sporting GJB2 genes, SLC26A4 genes and mtDNA,The deaf relevant mutational site is combined as:

    The mutation 299_300delAT and 235delC of the GJB2 genes, it is describedThe mutation 281C of SLC26A4 genes>

    T、

    919-2A>

    G、

    1226G>

    A、

    1229C>

    T、

    2027T>

    A、

    2168A>

    G and IVS15+5G>

    A,And the mutation 1494C of the mt DNA>

    T and 1555A>

    G,The amplimer includes the first primer and the second primer, is mutated for the 299_300delAT of the GJB2 genes, instituteIt is such as SEQ ID NO to state the first primer:

    Shown in 7, second primer is such as SEQ ID NO:

    Shown in 8, the extension primer isSuch as SEQ ID NO:

    Shown in 36;

    It is mutated for the 235delC of the GJB2 genes, first primer is such as SEQ ID NO:

    9Shown, second primer is such as SEQ ID NO:

    Shown in 10, the extension primer is such as SEQ ID NO:

    Shown in 37;

    For instituteState the 281C of SLC26A4 genes>

    T is mutated, and first primer is such as SEQ ID NO:

    Shown in 15, second primer be asSEQ ID NO:

    Shown in 16, the extension primer is such as SEQ ID NO:

    Shown in 40;

    For the 919-2A of the SLC26A4 genes>

    G is mutated, and first primer is such as SEQ ID NO:

    Shown in 19, second primer is such as SEQ ID NO:

    It is described shown in 20Extension primer is such as SEQ ID NO:

    Shown in 42;

    For the 1226G of the SLC26A4 genes>

    A is mutated, and first primer isSuch as SEQ ID NO:

    Shown in 21, second primer is such as SEQ ID NO:

    Shown in 22, the extension primer is such as SEQ IDNO:

    Shown in 44;

    For the 1229C of the SLC26A4 genes>

    T is mutated, and first primer is such as SEQ ID NO:

    Shown in 21,Second primer is such as SEQ ID NO:

    Shown in 22, the extension primer is such as SEQ ID NO:

    Shown in 45;

    For describedThe 2027T of SLC26A4 genes>

    A is mutated, and first primer is such as SEQ ID NO:

    Shown in 25, second primer is such as SEQID NO:

    Shown in 26, the extension primer is such as SEQ ID NO:

    Shown in 48;

    For the 2168A of the SLC26A4 genes>

    G dashes forwardBecome, first primer is such as SEQ ID NO:

    Shown in 27, second primer is such as SEQ ID NO:

    Shown in 28, the extensionPrimer is such as SEQ ID NO:

    Shown in 50;

    For the IVS15+5G of the SLC26A4 genes>

    A be mutated, first primer be asSEQ ID NO:

    Shown in 23, second primer is such as SEQ ID NO:

    Shown in 24, the extension primer is such as SEQ ID NO:

    Shown in 46;

    For the 1494C of the mt DNA genes>

    T is mutated, and first primer is such as SEQ ID NO:

    It is described shown in 29Second primer is such as SEQ ID NO:

    Shown in 30, the extension primer is such as SEQ ID NO:

    Shown in 51;

    And for the mtThe 1555A of DNA genes>

    G is mutated, and first primer is such as SEQ ID NO:

    Shown in 31, second primer is such as SEQ IDNO:

    Shown in 32, the extension primer is such as SEQ ID NO:

    Shown in 52.

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