Method for precise genetic testing by genomic hybridization

Abstract

Methods and apparatuses for selecting and arranging clinically relevant chromosomal loci allow an exemplary diagnostic array to simultaneously test for numerous genetic alterations that occur in many different parts of the human genome. Clinically irrelevant or ineffective loci are eliminated. One implementation increases reliability and accuracy by dividing the base-pair sequence of each chromosomal locus into segments and then assigning nucleic acid clones for comparative genomic hybridization to each different segment. The segments may overlap for increased resolution and control. Clones representing segments that are adjacent on a native chromosome are placed in non-adjacent target areas of the array to avoid interfering hybridization reactions. Arrangement motifs within an array may be redundantly repeated for high availability and increased reliability and accuracy of results. Techniques, hardware, software, logic engines, loci collections, and diagnostic arrays are described.

1 . A method for precise genetic testing, comprising:
 selecting clinically relevant chromosomal loci as nucleic acid hybridization targets, wherein each locus includes a base-pair sequence that is the site of a potential genetic alteration;   selecting multiple nucleic acid clones to represent each selected chromosomal locus, wherein the base-pair sequence is divided into multiple segments and each segment is mapped to by at least one of the multiple clones; and   subjecting each clone to comparative genomic hybridization in isolation from one or more clones that map to adjacent segments of the base-pair sequence.   
   
   
       2 . The method as recited in  claim 1 , wherein the base-pair sequence is divided into logically overlapping segments and each of the multiple clones map to one of the overlapping segments. 
   
   
       3 . The method as recited in  claim 1 , further comprising confirming that each clone maps uniquely to a single segment of the chromosomal locus and not to multiple chromosomal loci. 
   
   
       4 . The method as recited in  claim 3 , wherein the confirming is performed using fluorescence in situ hybridization. 
   
   
       5 . The method as recited in  claim 1 , further comprising discarding clones that ineffectively represent a segment of the chromosomal locus. 
   
   
       6 . The method as recited in  claim 5 , further comprising replacing a discarded clone with a clone that localizes to a respective segment. 
   
   
       7 . The method as recited in  claim 6 , further comprising verifying that a replacement clone localizes to a respective segment using fluorescence in situ hybridization. 
   
   
       8 . The method as recited in  claim 1 , wherein the base-pair sequence of the chromosomal locus is divided into a minimum of two overlapping segments. 
   
   
       9 . The method as recited in  claim 1 , wherein the selected chromosomal loci diagnose potential genetic alterations from multiple telomeric and pericentromeric regions of the human genome, wherein the selected chromosomal loci comprise 1p36.3, 1p12, 1q21, 1q44, 2p25.3, 2p11.2, 2q11.2, 2q37.3, 3p26.3, 3p11.2, 3q11.2, 3q29, 4p16.3, 4p12, 4q12, 4q35.2, 5p15.3, 5p12, 5q11.2, 5q35.3, 6p25.3, 6p11.2, 6q12, 6q27, 17p22.3, 7p11.2, 7q11.21, 7q36.3, 8p23.3, 8p11.2, 8q11.2, 8q24.3, 9p24.3, 9p11.2, 9q13, 9q34.3, 10p15.3, 10p11.21, 10q11.21, 10q26.3, 11p15.5, 11p11.2, 11q12, 11q25, 11p13.33, 12p11.21, 12q12, 12q24.33, 13q12.11, 13q34, 14q11.2, 14q32.33, 15q11.2, 15q26.3, 16p13.3, 16p11.2, 16q21.1, 16q24.3, 17p13.3, 17q11.2, 18p11.32, 18p11.21, 18q11.2, 18q23, 19p13.3, 19p12, 19q12, 19q13.43, 20p13, 20p11.21, 20q11.21, 20q13.33, 21q11.2, 21q22.3, 22q11.2, 22q13.3, Xp22.3, Xp11.22, Xq11.2, Xq28, YP11.3, Yp11.2, and YQ11.2; and
 wherein the selected chromosomal loci enable diagnosis of at least Holoprosencephaly 2, Nephronophthisis, Holoprosencephaly 6, Wolf-Hirschhorn, Cri-du-Chat, Saethre-Chotzen, Greig cephalopolysyndactyly, Williams syndrome, Holoprosencephaly 3, Langer-Giedion, Trichorhinophalangeal syndrome, Holoprosencephaly 7, DiGeorge syndrome II, Beckwith-Wiedemann, WAGR syndrome, Potocki-Shaffer syndrome, Noonan syndrome, Retinoblastoma/MR, Holoprosencephaly 5, Prader-Willi syndrome, Angelman syndrome, Rubinstein-Taybi, Tuberous sclerosis, Polycystic kidney disease, Miller-Dieker syndrome, Smith-Magenis syndrome, Duplication proximal 17, Neurofibromatosis I, Holoprosencephaly 4, Alagille syndrome, Holoprosencephaly 1, DiGeorge syndrome I, Steroid sulfatase deficiency, Microphthalmia with linear skin defects, Glycerol kinase deficiency, Adrenal Hypoplasia congenital, Duchenne muscular dystrophy, Pelizaeus-Merzbacher disease, and alterations of sex determining factor.   
   
   
       10 . The method as recited in  claim 1 , wherein the isolation comprises arranging clones that map to adjacent segments of the base-pair sequence in non-adjacent target areas of a comparative genomic hybridization array. 
   
   
       11 . A method for precise genetic testing, comprising:
 selecting clinically relevant chromosomal loci as nucleic acid hybridization targets, wherein each locus includes a base-pair sequence that is the site of a potential genetic alteration;   selecting multiple oligomers to represent each selected chromosomal locus, wherein the base-pair sequence is divided into multiple segments and each segment is mapped to by at least one of the oligomers; and   subjecting each oligomer to comparative genomic hybridization in isolation from one or more oligomers that map to adjacent segments of the base-pair sequence.   
   
   
       12 . The method as recited in  claim 11 , wherein the base-pair sequence is divided into logically overlapping segments and each of the multiple oligomers map to one of the overlapping segments. 
   
   
       13 . The method as recited in  claim 11 , further comprising confirming that each oligomer maps uniquely to a single segment of the chromosomal locus and not to multiple chromosomal loci. 
   
   
       14 . The method as recited in  claim 13 , wherein the confirming is performed using fluorescence in situ hybridization. 
   
   
       15 . The method as recited in  claim 11 , further comprising discarding oligomers that ineffectively represent a segment of the chromosomal locus. 
   
   
       16 . The method as recited in  claim 15 , further comprising replacing a discarded oligomer with an oligomer that localizes to a respective segment. 
   
   
       17 . The method as recited in  claim 16 , further comprising verifying that a replacement oligomer localizes to a respective segment using fluorescence in situ hybridization. 
   
   
       18 . The method as recited in  claim 11 , wherein the base-pair sequence of the chromosomal locus is divided into a minimum of two logically overlapping segments. 
   
   
       19 . The method as recited in  claim 11 , wherein the selected chromosomal loci diagnose potential genetic alterations from multiple telomeric and pericentromeric regions of the human genome, wherein the selected chromosomal loci comprise 1p36.3, 1p12, 1q21, 1q44, 2p25.3, 2p11.2, 2q11.2, 2q37.3, 3p26.3, 3p11.2, 3q11.2, 3q29, 4p16.3, 4p12, 4q12, 4q35.2, 5p15.3, 5p12, 5q11.2, 5q35.3, 6p25.3, 6p11.2, 6q12, 6q27, 17p22.3, 7p11.2, 7q11.21, 7q36.3, 8p23.3, 8p11.2, 8q11.2, 8q24.3, 9p24.3, 9p11.2, 9q13, 9q34.3, 10p15.3, 10p11.21, 10q11.21, 10q26.3, 11p15.5, 11p11.2, 11q12, 11q25, 11p13.33, 12p11.21, 12q12, 12q24.33, 13q12.11, 13q34, 14q11.2, 14q32.33, 15q11.2, 15q26.3, 16p13.3, 16p11.2, 16q21.1, 16q24.3, 17p13.3, 17q11.2, 18p11.32, 18p11.21, 18q11.2, 18q23, 19p13.3, 19p12, 19q12, 19q13.43, 20p13, 20p11.21, 20q11.21, 20q13.33, 21q11.2, 21q22.3, 22q11.2, 22q13.3, Xp22.3, Xp11.22, Xq11.2, Xq28, YP11.3, Yp11.2, and YQ11.2; and
 wherein the selected chromosomal loci enable diagnosis of at least Holoprosencephaly 2, Nephronophthisis, Holoprosencephaly 6, Wolf-Hirschhorn, Cri-du-Chat, Saethre-Chotzen, Greig cephalopolysyndactyly, Williams syndrome, Holoprosencephaly 3, Langer-Giedion, Trichorhinophalangeal syndrome, Holoprosencephaly 7, DiGeorge syndrome II, Beckwith-Wiedemann, WAGR syndrome, Potocki-Shaffer syndrome, Noonan syndrome, Retinoblastoma/MR, Holoprosencephaly 5, Prader-Willi syndrome, Angelman syndrome, Rubinstein-Taybi, Tuberous sclerosis, Polycystic kidney disease, Miller-Dieker syndrome, Smith-Magenis syndrome, Duplication proximal 17, Neurofibromatosis I, Holoprosencephaly 4, Alagille syndrome, Holoprosencephaly 1, DiGeorge syndrome I, Steroid sulfatase deficiency, Microphthalmia with linear skin defects, Glycerol kinase deficiency, Adrenal Hypoplasia congenital, Duchenne muscular dystrophy, Pelizaeus-Merzbacher disease, and alterations of sex determining factor.   
   
   
       20 . The method as recited in  claim 11 , wherein the isolation comprises arranging oligomers that map to adjacent segments of the base-pair sequence in non-adjacent target areas of a comparative genomic hybridization array.