AB INITIO GENERATION OF SINGLE COPY GENOMIC PROBES
First Claim
1. A method of producing a hybridization probe of a target reference complete genome sequence, wherein a single copy sequence is identified by a method of successive division of the target reference genome sequence into subintervals and comparison of the subintervals to the target reference sequence, said method comprising:
- determining a count of the number of times a subsequence of a first screened sequence occurs in the target reference genome sequence, said screened sequence being at least one subinterval of the target reference genome sequence obtained by division of the target reference genome sequence, whereinthe target reference genome sequence comprises the first screened sequence,the first screened sequence comprises at least two subsequences, anda single copy interval of the first screened sequence is identified as (i) a subsequence of the first screened sequence with a single subsequence occurrence in the target reference genome sequence, wherein an occurrence is defined by at least about 50 consecutive nucleotides of the subsequence having (i) at least about 60% homology to the target reference sequence;
(ii) at least about 70% homology to the target reference sequence;
or (iii) at least about 80% homology to the target reference sequence, or (ii) a group of contiguous subsequences of the first screened sequence, each member being a single subsequence occurrence in the target reference genome sequence, wherein an occurrence is defined by at least about 50 consecutive nucleotides of the group of contiguous subsequences having (i) at least about 60% homology to the target reference sequence;
(ii) at least about 70% homology to the target reference sequence;
or (iii) at least about 80% homology to the target reference sequence;
determining a count of the number of times a subsequence of a second screened sequence occurs in the target reference genome sequence, said screened sequence being at least one subinterval of the target reference genome sequence, whereinthe second screened sequence comprises a single copy interval of the first screened sequence;
the second screened sequence overlaps the single copy interval of the first screened sequence;
the subsequences of the second screened sequence are either (i) consecutive non-overlapping subintervals of the second screened sequence or (ii) overlapping non-identical subintervals of the second screened sequence; and
(4) a single copy interval of the second screened sequence is identified as (i) a subsequence of the second screened sequence with a single subsequence occurrence in the target reference genome sequence, wherein an occurrence is defined by at least about 50 consecutive nucleotides of the subsequence having (i) at least about 60% homology to the target reference sequence;
(ii) at least about 70% homology to the target reference sequence;
or (iii) at least about 80% homology to the target reference sequence, or (ii) a group of contiguous subsequences of the second screened sequence, each member being a single subsequence occurrence in the target reference genome sequence, wherein an occurrence is defined by at least about 50 consecutive nucleotides of the group of contiguous subsequences having (i) at least about 60% homology to the target reference sequence;
(ii) at least about 70% homology to the target reference sequence;
or (iii) at least about 80% homology to the target reference sequence; and
(C) identifying a single copy interval and at least one contiguous divergent repetitive interval of the target reference sequence wherein at least one subsequence in the target sequence contains a divergent repetitive element suitable for use as a probe that hybridizes to a single location in the target genome, wherein said divergent repetitive element is washed under conditions that eliminate cross-hybridization to other target sequences in the genome.
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Abstract
Single copy sequences suitable for use as DNA probes can be defined by computational analysis of genomic sequences. The present invention provides an ab initio method for identification of single copy sequences for use as probes which obviates the need to compare genomic sequences with existing catalogs of repetitive sequences. By dividing a target reference sequence into a series of shorter contiguous sequence windows and comparing these sequences with the reference genome sequence, one can identify single copy sequences in a genome. Probes can then be designed and produced from these single copy intervals.
146 Citations
18 Claims
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1. A method of producing a hybridization probe of a target reference complete genome sequence, wherein a single copy sequence is identified by a method of successive division of the target reference genome sequence into subintervals and comparison of the subintervals to the target reference sequence, said method comprising:
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determining a count of the number of times a subsequence of a first screened sequence occurs in the target reference genome sequence, said screened sequence being at least one subinterval of the target reference genome sequence obtained by division of the target reference genome sequence, wherein the target reference genome sequence comprises the first screened sequence, the first screened sequence comprises at least two subsequences, and a single copy interval of the first screened sequence is identified as (i) a subsequence of the first screened sequence with a single subsequence occurrence in the target reference genome sequence, wherein an occurrence is defined by at least about 50 consecutive nucleotides of the subsequence having (i) at least about 60% homology to the target reference sequence;
(ii) at least about 70% homology to the target reference sequence;
or (iii) at least about 80% homology to the target reference sequence, or (ii) a group of contiguous subsequences of the first screened sequence, each member being a single subsequence occurrence in the target reference genome sequence, wherein an occurrence is defined by at least about 50 consecutive nucleotides of the group of contiguous subsequences having (i) at least about 60% homology to the target reference sequence;
(ii) at least about 70% homology to the target reference sequence;
or (iii) at least about 80% homology to the target reference sequence;determining a count of the number of times a subsequence of a second screened sequence occurs in the target reference genome sequence, said screened sequence being at least one subinterval of the target reference genome sequence, wherein the second screened sequence comprises a single copy interval of the first screened sequence; the second screened sequence overlaps the single copy interval of the first screened sequence; the subsequences of the second screened sequence are either (i) consecutive non-overlapping subintervals of the second screened sequence or (ii) overlapping non-identical subintervals of the second screened sequence; and
(4) a single copy interval of the second screened sequence is identified as (i) a subsequence of the second screened sequence with a single subsequence occurrence in the target reference genome sequence, wherein an occurrence is defined by at least about 50 consecutive nucleotides of the subsequence having (i) at least about 60% homology to the target reference sequence;
(ii) at least about 70% homology to the target reference sequence;
or (iii) at least about 80% homology to the target reference sequence, or (ii) a group of contiguous subsequences of the second screened sequence, each member being a single subsequence occurrence in the target reference genome sequence, wherein an occurrence is defined by at least about 50 consecutive nucleotides of the group of contiguous subsequences having (i) at least about 60% homology to the target reference sequence;
(ii) at least about 70% homology to the target reference sequence;
or (iii) at least about 80% homology to the target reference sequence; and(C) identifying a single copy interval and at least one contiguous divergent repetitive interval of the target reference sequence wherein at least one subsequence in the target sequence contains a divergent repetitive element suitable for use as a probe that hybridizes to a single location in the target genome, wherein said divergent repetitive element is washed under conditions that eliminate cross-hybridization to other target sequences in the genome. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16)
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17. A method of producing a hybridization probe of a target reference complete genome sequence, wherein a single copy sequence is identified by a method of successive division of the target reference genome sequence into subintervals and comparison of the subintervals to the target reference sequence, said method comprising:
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determining a count of the number of times a subsequence of a first screened sequence occurs in the target reference genome sequence, said screened sequence being at least one subinterval of the target reference genome sequence obtained by division of the target reference genome sequence, wherein the target reference genome sequence comprises the first screened sequence, the first screened sequence comprises at least two subsequences, and a single copy interval of the first screened sequence is identified as (i) a subsequence of the first screened sequence with a single subsequence occurrence in the target reference genome sequence, wherein an occurrence is defined by at least about 50 consecutive nucleotides of the subsequence having (i) at least about 60% homology to the target reference sequence;
(ii) at least about 70% homology to the target reference sequence;
or (iii) at least about 80% homology to the target reference sequence, or (ii) a group of contiguous subsequences of the first screened sequence, each member being a single subsequence occurrence in the target reference genome sequence, wherein an occurrence is defined by at least about 50 consecutive nucleotides of the group of contiguous subsequences having (i) at least about 60% homology to the target reference sequence;
(ii) at least about 70% homology to the target reference sequence;
or (iii) at least about 80% homology to the target reference sequence;determining a count of the number of times a subsequence of a second screened sequence occurs in the target reference genome sequence, said screened sequence being at least one subinterval of the target reference genome sequence, wherein the second screened sequence comprises a single copy interval of the first screened sequence; the second screened sequence overlaps the single copy interval of the first screened sequence; the subsequences of the second screened sequence are either (i) consecutive non-overlapping subintervals of the second screened sequence or (ii) overlapping non-identical subintervals of the second screened sequence; and
(4) a single copy interval of the second screened sequence is identified as (i) a subsequence of the second screened sequence with a single subsequence occurrence in the target reference genome sequence, wherein an occurrence is defined by at least about 50 consecutive nucleotides of the subsequence having (i) at least about 60% homology to the target reference sequence;
(ii) at least about 70% homology to the target reference sequence;
or (iii) at least about 80% homology to the target reference sequence, or (ii) a group of contiguous subsequences of the second screened sequence, each member being a single subsequence occurrence in the target reference genome sequence, wherein an occurrence is defined by at least about 50 consecutive nucleotides of the group of contiguous subsequences having (i) at least about 60% homology to the target reference sequence;
(ii) at least about 70% homology to the target reference sequence;
or (iii) at least about 80% homology to the target reference sequence; and(C) identifying a single copy interval and at least one contiguous divergent repetitive interval of the target reference sequence wherein at least one subsequence in the target sequence contains a divergent repetitive element suitable for use as a probe that hybridizes to a single location in the target genome, wherein said divergent repetitive element is washed under conditions that eliminate cross-hybridization to other target sequences in the genome, where such conditions comprise washing the hybridized probe in a solution of 0.1×
SSC (15 mM NaCl and 1.5 mM Na3C6H5O7) at a temperature exceeding 42 degrees Celsius.
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18. A method of producing a hybridization probe of a target reference complete genome sequence, wherein a single copy sequence is identified by a method of successive division of the target reference genome sequence into subintervals and comparison of the subintervals to the target reference sequence, said method comprising:
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determining a count of the number of times a subsequence of a first screened sequence occurs in the target reference genome sequence, said screened sequence being at least one subinterval of the target reference genome sequence obtained by division of the target reference genome sequence, wherein the target reference genome sequence comprises the first screened sequence, the first screened sequence comprises at least two subsequences, and a single copy interval of the first screened sequence is identified as (i) a subsequence of the first screened sequence with a single subsequence occurrence in the target reference genome sequence, wherein an occurrence is defined by at least about 50 consecutive nucleotides of the subsequence having (i) at least about 60% homology to the target reference sequence;
(ii) at least about 70% homology to the target reference sequence;
or (iii) at least about 80% homology to the target reference sequence, or (ii) a group of contiguous subsequences of the first screened sequence, each member being a single subsequence occurrence in the target reference genome sequence, wherein an occurrence is defined by at least about 50 consecutive nucleotides of the group of contiguous subsequences having (i) at least about 60% homology to the target reference sequence;
(ii) at least about 70% homology to the target reference sequence;
or (iii) at least about 80% homology to the target reference sequence;determining a count of the number of times a subsequence of a second screened sequence occurs in the target reference genome sequence, said screened sequence being at least one subinterval of the target reference genome sequence, wherein the second screened sequence comprises a single copy interval of the first screened sequence; the second screened sequence overlaps the single copy interval of the first screened sequence; the subsequences of the second screened sequence are either (i) consecutive non-overlapping subintervals of the second screened sequence or (ii) overlapping non-identical subintervals of the second screened sequence; and
(4) a single copy interval of the second screened sequence is identified as (i) a subsequence of the second screened sequence with a single subsequence occurrence in the target reference genome sequence, wherein an occurrence is defined by at least about 50 consecutive nucleotides of the subsequence having (i) at least about 60% homology to the target reference sequence;
(ii) at least about 70% homology to the target reference sequence;
or (iii) at least about 80% homology to the target reference sequence, or (ii) a group of contiguous subsequences of the second screened sequence, each member being a single subsequence occurrence in the target reference genome sequence, wherein an occurrence is defined by at least about 50 consecutive nucleotides of the group of contiguous subsequences having (i) at least about 60% homology to the target reference sequence;
(ii) at least about 70% homology to the target reference sequence;
or (iii) at least about 80% homology to the target reference sequence; and(C) identifying a single copy interval and at least one contiguous divergent repetitive interval of the target reference sequence wherein at least one subsequence in the target sequence contains a divergent repetitive element suitable for use as a probe that hybridizes to a single location in the target genome, wherein said divergent repetitive element is washed under conditions that eliminate cross-hybridization to other target sequences in the genome, where such conditions comprise washing the hybridized probe in a solution of 0.2×
SSC (30 mM NaCl and 3 mM Na3C6H5O7) at a temperature exceeding 37 degrees Celsius.
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Specification