Assay for human DNA for gender determination

• US 7,432,362 B2
• Filed: 05/10/2006
• Issued: 10/07/2008
• Est. Priority Date: 09/30/2003
• Status: Expired due to Term

• Current Assignee: Reliagene Technologies Inc. (Laboratory Corporation Of America Holdings), Life Genetics Lab, LLC, Board of Supervisors of Louisiana State University Agricultural & Mechanical College (Louisiana State University System)
• Original Assignee: Reliagene Technologies Inc. (Laboratory Corporation Of America Holdings), Board of Supervisors of Louisiana State University Agricultural & Mechanical College (Louisiana State University System)
• Inventors: Batzer, Mark A., Sinha, Sudhir K., Hedges, Dale J.
• Primary Examiner(s): Horlick; Kenneth R.
• Assistant Examiner(s): Babic; Christopher M.
• Application Number:
  US11/431,030
• Publication Number:
  US 20060199217A1
• Time in Patent Office: 881 Days
• Field of Search: None
• US Class Current: 536/23.1
• CPC Class Codes: C12Q 1/6876 Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
• CPC Class Codes: C12Q 1/6879 for sex determination
• CPC Class Codes: C12Q 2600/156 Polymorphic or mutational markers
• CPC Class Codes: Y10S 435/975 Kit

Abstract

A method for determining gender from a human DNA sample. The loci of Alu element insertion is selected, amplified and evaluated in terms of size of the fragment. The gender assay utilizes AluSTXa for the X chromosome, AluSTYa for the Y chromosome, or both AluSTXa and AluSTYa, to reduce the possibility of error to a negligible quantity. The inserted chromosome yields a large fragment when the homologous region is amplified. The males are distinguished as having two DNA amplicons present, while females have only a single amplicon. The kit adapted for carrying out the method includes a pair of primers to amplify the locus and optionally polymerase chain reaction regents.

Claims

1. A primer pair for a sex determination of a human DNA sample, said primer pair selected from the group consisting of the following primer pairs:

a primer pair comprising;

a forward primer having the sequence consisting of;

5′

-ACTGCAGTCCGCAGTCCGG-3′

(SEQ ID NO;

     1); and

a reverse primer having the sequence consisting of;

5′

-CCACTGACATATTATCAGCCTTATTAC-3′

(SEQ ID NO;

     2);

a primer pair comprising;

a forward primer having the sequence consisting of;

5′

-TGAAGAAATTCAGTTCATAGCTTGT-3′

(SEQ ID NO;

     3); and

a reverse primer having the sequence consisting of;

5′

-CAGGAGATCCTGAGATTATGTGG-3′

(SEQ ID NO;

     4); and

a primer pair comprising;

a forward primer consisting of;

5′

-AAACATGGCATGGGCCTCT-3′

(SEQ ID NO;

     5); and

a reverse primer consisting of;

5′

-TTTTAAAGCCACAGACCTCTCTCT-3′

(SEQ ID NO;

     6).

2. The primer pair of claim 1, wherein each of the primer pair has the sequence selected from the group consisting of:

3. The primer pair of claim 1, wherein each of the primer pair has the sequence selected from the group consisting of:

4. The primer pair of claim 1, wherein each primer of the primer pair has the sequence selected from the group consisting of:

5. A kit for determining gender from a sample, comprising:

a pair of primers capable of amplifying a selected locus of a human DNA in an amplification reaction, said selected locus being in a non-combining X-Y homologous region and selected from the group consisting of a locus in AluSTYa on the Y chromosome and a locus in AluSTXa on the X chromosome, said region containing a monomorphic Alu insertion in one of the X chromosome and the Y chromosome, said pair of primers selected from the group consisting of the following primer pairs;

a primer pair comprising a forward primer consisting of 5′

-ACTGCAGTCCGCAGTCCGG-3′

(SEQ ID NO;

     1) and a reverse primer having the sequence consisting of 5′

-CCACTGACATATTATCAGCCTTATTAC-3′

(SEQ ID NO;

     2);

a primer pair comprising a forward primer consisting of 5′

-TGAAGAAATTCAGTTCATAGCTTGT-3′

(SEQ ID NO;

     3) and a reverse primer consisting of 5′

- CAGGAGATCCTGAGATTATGTGG-3′

(SEQ ID NO;

     4); and

a primer pair comprising a forward primer having the sequence consisting of 5′

-AAACATGGCATGGGCCTCT-3′

(SEQ ID NO;

     5) and a reverse primer having the sequence consisting of 5′

-TTTTAAAGCCACAGACCTCTCTCT-3′

(SEQ ID NO;

     6); and

a polymerase chain reaction reagent comprising a polymerase catalyzing the amplification of the selected locus.

6. The kit of claim 5, wherein the selected locus is AluSTYa locus on the Y chromosome.

7. The kit of claim 5, wherein each of the primer pair has the sequence selected from the group consisting of:

8. The kit of claim 5, wherein the selected locus is AluSTXa locus on the X chromosome.

9. The kit of claim 5, wherein each of the primer pair has the sequence selected from the group consisting of:

10. The kit of claim 5, wherein the selected locus comprises AluSTYa locus and AluSTXa locus.

11. The kit of claim 5, wherein said pair of primers comprises:

a first primer pair comprising a forward primer consisting of 5′

-TGAAGAAATTCAGTTCATAGCTTGT-3′

(SEQ ID NO;

     3) and a reverse primer consisting of 5′

-CAGGAGATCCTGAGATTATGTGG-3′

(SEQ ID NO;

     4); and

a second primer pair comprising a forward primer consisting of 5′

-ACTGCAGTCCGCAGTCCGG-3′

(SEQ ID NO;

     1) and a reverse primer consisting of 5′

-CCACTGACATATTATCAGCCTTATTAC-3′

(SEQ ID NO;

     2).

12. A kit for determining gender from a sample, comprising:

a pair of primers comprising a forward primer consisting of 5′

-AAACATGGCATGGGCCTCT-3′

(SEQ ID NO;

     5) and a reverse primer consisting of 5′

-TTTAAAGCCACAGACCTCTCTCT-3′

(SEQ ID NO;

     6); and

a polymerase chain reaction reagent comprising a polymerase catalyzing the amplification of the alleles.