Cystic fibrosis transmembrane conductance regulator gene mutations

• US 20070161013A1
• Filed: 08/18/2006
• Published: 07/12/2007
• Est. Priority Date: 08/18/2005
• Status: Active Grant

• Current Assignee: Quest Diagnostics Investments Incorporated (Quest Diagnostics Incorporated)
• Original Assignee: Quest Diagnostics Incorporated
• Inventors: Hantash, Feras
• Granted Patent: US 7,838,646 B2
• Time in Patent Office: Days
• US Class Current: 435/6
• CPC Class Codes: C12Q 1/6883 for diseases caused by alterations of genetic material
• CPC Class Codes: C12Q 2600/156 Polymorphic or mutational markers
• CPC Class Codes: C12Q 2600/16 Primer sets for multiplex assays
• CPC Class Codes: C12Q 2600/172 Haplotypes

Abstract

The present invention provides novel mutations of the CFTR gene related to cystic fibrosis or to conditions associated with cystic fibrosis. The mutations include duplication of exons including duplication of exons 6b through 10. Methods of identifying if an individual contains the exons 6b through 10 duplication are provided as well as nucleic acid fragments that contain the junction site of the duplicated segment. The detection of additional mutations in the CFTR gene are also provided.

Claims

1. A method of detecting if a CFTR gene in nucleic acid from an individual has a duplication of exons 6b through 10 by detecting the duplication of at least one portion of exons 6b through 10.

2. The method of claim 1 wherein the individual is tested to determine if the individual is a cystic fibrosis (CF) carrier or a CF patient.

3. The method of claim 1 wherein the individual is being tested to determine if the individual is predisposed to cystic fibrosis or a condition associated with cystic fibrosis.

4. The method of claim 1 wherein the individual is being tested to counsel on the likelihood of having an offspring afflicted with cystic fibrosis or a condition associated with cystic fibrosis.

5. The method of claim 1 wherein both alleles in the individual are evaluated for said duplication.

6. The method of claim 1 wherein genomic DNA is tested for the presence of said CFTR gene duplication.

7. The method of claim 1 wherein a cDNA copy of the CFTR gene coding sequence is tested for the presence of said duplication.

8. The method of claim 1 wherein said mutation is detected by:

(a) amplifying sequence from one or more of exon 6b, 7, 8, 9, and 10 from nucleic acid from said individual;

(b) identifying the amplified exon sequence; and

(c) comparing the amount of each amplified exon sequence versus that similarly amplified for a normal CFTR gene, wherein a substantial increase in the amount of amplified exon sequence observed versus that for a normal cystic fibrosis transmembrane regulatory gene indicates a duplication of exon 6b-10.

9. The method of claim 8 wherein sequence is amplified using oligonucleotide primer pairs that hybridize upstream and downstream of said amplified sequence.

10. The method of claim 9 wherein at least one primer of each said primer pair is labeled with a detectable moiety.

11. The method of claim 1 wherein sequence from the CFTR gene is amplified and the amplified sequence is tested for the presence of said duplication.

12. The method of claim 11 wherein said nucleic acid assay comprises:

amplifying across a junction site between exons 10 and 6b resulting from the 6b-10 duplication and detecting the amplified product.

13. The method of claim 12 wherein said junction site is IVS10+2,987/IVS6a+415.

14. The method of claim 1 wherein the duplication is detected using multiplex ligation-dependent probe amplification or multiplex amplifiable probe hybridization.

15. The method of claim 1 wherein the 6b-10 duplication is detected by Southern blotting.

16. A nucleic acid fragment of about 26 kb or less, comprising all or a portion of a intron 10 to intron 6a junction formed as a result of an exon 6b to 10 duplication of the CFTR gene, wherein the junction includes a sequence with at least nine nucleotides directly adjoining each side of the junction.

17. A nucleic acid fragment of claim 16 wherein the sequence of the junction with at least nine nucleotides directly adjoining each side of the junction is

18. A nucleic acid fragment of claim 16 wherein the sequence of the junction with at least nine nucleotides directly adjoining each side of the junction is

19. A nucleic acid fragment of claim 16 wherein the sequence of the junction with at least nine nucleotides directly adjoining each side of the junction is