Systems and methods for designing and ordering polynucleotides

• US 20070043516A1
• Filed: 08/16/2005
• Published: 02/22/2007
• Est. Priority Date: 08/16/2005
• Status: Active Grant

• Current Assignee: DNA Twopointo Incorporated
• Original Assignee: DNA Twopointo Incorporated
• Inventors: Govindarajan, Sridhar, Gustafsson, Claes, Minshull, Jeremy, Ness, Jon, Villalobos, Alan
• Granted Patent: US 7,805,252 B2
• Time in Patent Office: Days
• US Class Current: 702/20
• CPC Class Codes: G16B 20/00 ICT specially adapted for functional genomics or proteomics, e.g. genotype-phenotype associations
• CPC Class Codes: G16B 20/30 Detection of binding sites or motifs
• CPC Class Codes: G16B 20/50 Mutagenesis
• CPC Class Codes: G16B 25/00 ICT specially adapted for hybridisation; ICT specially adapted for gene or protein expression
• CPC Class Codes: G16B 25/20 Polymerase chain reaction [PCR]; Primer or probe design; Probe optimisation
• CPC Class Codes: G16B 45/00 ICT specially adapted for bioinformatics-related data visualisation, e.g. displaying of maps or networks

Abstract

Computer systems, computer program products and methods for designing oligonucleotides are provided. A set of sequence elements is defined. Each sequence element represents an amino acid sequence segment or a nucleic acid sequence segment. The set of sequence elements collectively represent a design nucleic acid sequence. The set of sequence elements are displayed as a plurality icons in a linear or a near linear arrangement such that each respective icon in the plurality of icons uniquely represents a corresponding sequence element in the set of sequence elements. In this representation, neighboring icons in the plurality of icons represent neighboring sequence elements in the set of sequence elements. Each respective icon in the plurality of icons depicts a directional property for the corresponding sequence element in the set of sequence elements. An oligonucleotide selection module is used to identify oligonucleotides in the design nucleic acid sequence.

Claims

1. A computer program product for use in conjunction with a computer system, the computer program product comprising a computer readable storage medium and a computer program mechanism embedded therein, the computer program mechanism comprising:

instructions for representing a set of sequence elements, each sequence element representing an amino acid sequence segment or a nucleic acid sequence segment, wherein the set of sequence elements are collectively represented by a design nucleic acid sequence, wherein

said instructions for representing said set of sequence elements comprise instructions for displaying a plurality of icons in a linear or a near linear arrangement, each respective icon in said plurality of icons uniquely representing a corresponding sequence element in said set of sequence elements such that neighboring icons in said plurality of icons represent neighboring sequence elements in said plurality of sequence elements, and

each said respective icon in said plurality of icons depicts a directional property for the corresponding sequence element in said set of sequence elements.

2. The computer program product of claim 1, wherein said directional property for a corresponding sequence element in said set of sequence elements is a translation direction or a transcription direction.

3. The computer program product of claim 1, wherein

said instructions for displaying a set of icons further comprise instructions for displaying an icon in said plurality of icons in an icon view or a sequence view, wherein,

when said icon is displayed in the icon view, a graphical depiction of the sequence element represented by said icon is displayed, and

when said icon is displayed in the sequence view, a sequence represented by said icon is displayed.

4. The computer program product of claim 3, wherein, when said icon is displayed in said sequence view, a name of the sequence element represented by the icon is displayed above said sequence.

5. The computer program product of claim 3, wherein, when said icon is displayed in said sequence view, a start position and an end position of the sequence element that said icon represents in said design nucleic acid sequence is displayed.

6. The computer program product of claim 3, wherein the sequence represented by said icon is an amino acid sequence and, for each respective amino acid in said amino acid sequence, each codon corresponding to said respective amino acid is displayed below the respective amino acid.

7. The computer program product of claim 3, wherein the sequence represented by said icon is an amino acid sequence.

8. The computer program product of claim 3, wherein the sequence represented by said icon is a nucleic acid sequence.

9. The computer program product of claim 1, the computer program mechanism further comprising:

a library of sequence elements;

instructions for depicting said library of sequence elements; and

instructions for permitting a user to drag a copy of a sequence element in said library of sequence elements onto a panel, thereby incorporating said sequence element into said set of sequence elements.

10. The computer program product of claim 9, the computer program mechanism further comprising:

instructions for permitting a user to modify a sequence element in said set of sequence elements; and

instructions for permitting a user to add a sequence element from said set of sequence elements to said library of sequence elements.

11. The computer program product of claim 9, wherein the library of sequence elements are organized in a hierarchical tree that is graphically displayed.

12. The computer program product of claim 9, wherein the library of sequence elements are organized in a hierarchical tree that is graphically displayed, and wherein said hierarchical tree is divided into a first portion representing regulatory elements, a second portion representing expressed elements, and a third portion representing cloning elements.

13. The computer program product of claim 12, wherein the portion of said hierarchical tree representing regulatory elements is further divided into a subportion representing sequence elements that are transcriptional elements and a subportion representing sequence elements that are translational elements.

14. The computer program product of claim 13, wherein the portion of said hierarchical tree representing transcriptional elements is further divided into one or more of the following subportions:

a subportion representing sequence elements that are enhancers;

a subportion representing sequence elements that are promoters;

a subportion representing sequence elements that are operators;

a subportion representing sequence elements that are terminators; and

a subportion representing sequence elements that are polyadenylation signals.

15. The computer program product of claim 13, wherein the portion of said hierarchical tree representing translational elements is further divided into one or more of the following subportions:

a subportion representing sequence elements that are 5′ and

3′

untranslated regions;

a subportion representing sequence elements that are ribosome binding sites;

a subportion representing sequence elements that are initiation AUG contexts; and

a subportion representing sequence elements that are termination codons.

16. The computer program product of claim 12, wherein the portion of said hierarchical tree representing expressed elements is further divided into one or more of the following subportions:

a subportion representing sequence elements that are peptide fusion tags;

a subportion representing sequence elements that are protease cleavage sites;

a subportion representing sequence elements that are solubility or fusion tags; and

a subportion representing sequence elements that are secretion signals.

17. The computer program product of claim 16, wherein the portion of said hierarchical tree representing expressed elements is further divided according to organism of origin.

18. The computer program product of claim 12, wherein the portion of said hierarchical tree representing cloning elements is further divided into one or more of the following subportions:

a subportion representing sequence elements that are recombinase recognition sequences; and

a subportion representing sequence elements that are restriction enzyme recognition sequences.

19. The computer program product of claim 1, the computer program mechanism further comprising

instructions for depicting an empty sequence element;

and

instructions for permitting a user to drag a copy of the empty sequence element onto a panel, thereby incorporating said empty sequence element into said set of sequence elements, wherein, when said user drags said copy of the empty sequence element onto said panel said instructions for permitting a user further comprise instructions for prompting said user for an amino acid sequence, a nucleic acid sequence, or an open reading frame.

20. The computer program product of claim 1, wherein each sequence element in said set of sequence elements is an amino acid element, a DNA element, or an open reading frame element.

21. The computer program product of claim 20, the computer program product further comprising instructions for back-translating a sequence element in said set of sequence elements that is an amino acid element or an open reading frame element into said design nucleic acid sequence.

22. The computer program product of claim 21, wherein the sequence element is an amino acid sequence and the instructions for back-translating produce the corresponding nucleic acid sequence as a function of (i) the amino acid sequence and (ii) common codon use in a designated species.

23. The computer program product of claim 21, wherein

the instructions for back-translating further comprise instructions for avoiding the generation of one or more restriction enzyme recognition sequences in said design nucleic acid sequence;

and wherein

the computer program mechanism further comprises instructions for receiving an identity of the one or more restriction enzyme recognition sequences to be avoided from a user.

24. The computer program product of claim 21, wherein the instructions for back-translating further comprise instructions for considering one or more criteria when back-translating, the one or more criteria selected from the group consisting of:

(i) minimization of a repeated nucleic acid sequence in the design nucleic acid sequence;

(ii) avoidance of a predetermined nucleic acid sequence in the design nucleic acid sequence;

(iii) minimization of a secondary structure in the design nucleic acid sequence;

(iv) minimization of sequence identity of the design nucleic acid sequence with respect to a reference sequence or maximization of sequence identity with respect to said reference sequence;

(v) avoidance of an enzyme recognition sequence in the design nucleic acid sequence;

(vi) selection of a codon based on codon frequency specified by a codon table;

(vii) elimination of a methylation site that would inhibit the action of an enzyme in the design nucleic acid sequence; and

(viii) avoidance of a first subsequence in the design nucleic acid sequence that has an annealing temperature with a second subsequence in the design nucleic acid sequence that is above a predetermined value.

25. The computer program product of claim 21, the computer program product further comprising instructions for fixing and unfixing the sequence element, wherein,

when the sequence element is fixed, it cannot be subjected to back-translation optimization;

and

when the sequence element is unfixed, it can be further subjected to back-translation optimization.

26. The computer program product of claim 20, wherein the computer program mechanism further comprises:

instructions for independently toggling an open reading frame element in said set of sequence elements between a fixed state and an unfixed state; and

instructions for back-translating a sequence element in said set of sequence elements that is an open reading frame element in an unfixed state into the design nucleic acid sequence.

27. The computer program product of claim 26, wherein

the instructions for back-translating further comprise instructions for avoiding the generation of one or more restriction enzyme recognition sequences in said design nucleic acid sequence;

and wherein

the computer program mechanism further comprises instructions for receiving an identity of the one or more restriction enzyme recognition sequences to be avoided from a user.

28. The computer program product of claim 26, wherein the instructions for back-translating further comprise instructions for considering one or more criteria when back-translating, the one or more criteria selected from the group consisting of:

(i) minimization of a repeated nucleic acid sequence in the design nucleic acid sequence;

(ii) avoidance of a predetermined nucleic acid sequence in the design nucleic acid sequence;

(iii) minimization of a secondary structure in the design nucleic acid sequence;

(iv) minimization of sequence identity of the design nucleic acid sequence with respect to a reference sequence or maximization of sequence identity with respect to said reference sequence;

(v) avoidance of an enzyme recognition sequence in the design nucleic acid sequence;

(vi) selection of a codon based on codon frequency specified by a codon table;

(vii) elimination of a methylation site that would inhibit the action of an enzyme in the design nucleic acid sequence; and

(viii) avoidance of a first subsequence in the design nucleic acid sequence that has an annealing temperature with a second subsequence in the design nucleic acid sequence that is above a predetermined value.

29. The computer program product of claim 26, the computer program product further comprising instructions for fixing and unfixing the sequence element, wherein,

when the sequence element is fixed, it cannot be subjected to further back-translation optimization;

and

when the sequence element is unfixed, it can be further subjected to back-translation optimization.

30. The computer program product of claim 1, wherein said linear or near linear arrangement is broken into a plurality of horizontal rows.

31. The computer program product of claim 1, wherein the computer program mechanism further comprises:

instructions for permitting a user to rearrange an order of said sequence elements in said linear or near linear arrangement thereby causing a corresponding change in said design nucleic acid sequence.

32. The computer program product of claim 1, wherein the computer program mechanism further comprises:

instructions for permitting a user to change an attribute of an icon in said plurality of icons.

33. The computer program product of claim 32, wherein the attribute is a name of said icon, a color of said icon, a size of said icon, a resolution of said icon, a sequence associated with said icon or a name associated with said icon.

34. The computer program product of claim 1, wherein a sequence element in said set of sequence elements is a DNA element, wherein said instructions for displaying a set of icons further comprise instructions for displaying a first icon in said plurality of icons representing said DNA element in an icon view or a sequence view, wherein,

when said first icon is displayed in the icon view, a graphical depiction of the sequence element represented by said first icon is displayed, and

when said first icon is displayed in the sequence view, an nucleic acid sequence represented by said first icon is displayed and all six reading frames of said nucleic acid sequence represented by said icon is displayed.

35. The computer program product of claim 1, wherein a sequence element in said set of sequence elements is an amino acid element, the computer program product further comprising:

instructions for back-translating said amino acid element to said design nucleic acid sequence; and

wherein

said instructions for displaying a set of icons further comprise instructions for displaying an icon in said plurality of icons representing said amino acid element in an icon view or a sequence view, wherein,

when said icon is displayed in the icon view, a graphical depiction of the sequence element represented by said icon is displayed, and

when said icon is displayed in the sequence view, an amino acid sequence represented by said icon is displayed and, for each respective amino acid in said amino acid sequence, each possible codon for said respective amino acid is displayed below the respective amino acid.

36. The computer program product of claim 35, wherein codons for each respective amino acid in said amino acid sequence are ranked in the sequence view in accordance with frequencies found for such codons in a codon bias table.

37. The computer program product of claim 36, the computer program product further comprising instructions for obtaining said codon bias table from among a plurality of codon bias tables, wherein said codon bias table indicates a frequency for each possible codon encoding a naturally occurring amino acid.

38. The computer program product of claim 37, wherein said frequency for each possible codon encoding a naturally occurring amino acid is the frequency of occurrence of each possible codon encoding a naturally occurring amino acid in a species corresponding to said codon bias table.

39. The computer program product of claim 37, wherein said frequency for each possible codon encoding a naturally occurring amino acid is calculated from the frequency of occurrence of each possible codon encoding a naturally occurring amino acid in two or more species corresponding to said codon bias table.

40. The computer program product of claim 36, the computer program product further comprising:

instructions for setting a back-translation threshold; and

wherein

said instructions for back-translating include instructions for excluding codons in said corresponding nucleic acid sequence that are below said back-translation threshold in a codon bias table.

41. The computer program product of claim 40, the computer program product further comprising:

instructions for displaying codons that fall below said back-translation threshold in a first color; and

instructions for displaying codons that are above said back-translation threshold in a second color.

42. The computer program product of claim 1, the computer program product further comprising:

instructions for displaying a restriction site analysis box comprising a plurality of restrictions site names and, for each restriction site name in the plurality of restriction site names, the corresponding restriction sequence; and

instructions for indicating positions of one or more restriction sites in the plurality of restriction sites in said set of sequence elements that are displayed as a plurality of icons in said linear or said near linear arrangement.

43. The computer program product of claim 42, wherein a sequence element in said set of sequence elements comprises an amino acid sequence or an open reading frame and wherein said instructions for indicating positions of one or more restriction sites comprises instructions for indicating a position where a restriction site could occur in said amino acid sequence or said open reading frame without changing said amino acid sequence or said open reading frame.

44. The computer program product of claim 42, the computer program product further comprising:

instructions for adding or removing a restriction site name to said plurality of restriction site names; and

instructions for selecting and deselecting a restriction site name in said plurality of restriction site names, wherein, when the restriction site name is selected the position of each restriction site in said set of sequence elements corresponding to said restriction site name is displayed.

45. The computer program product of claim 1, wherein a sequence element in said set of sequence elements is an open reading frame element, the computer program product further comprising:

instructions for setting a codon usage threshold; and

wherein

said instructions for displaying a set of icons further comprise instructions for displaying a first icon in said plurality of icons representing said open reading frame element in an icon view or a sequence view, wherein,

when said first icon is displayed in the icon view, a graphical depiction of the sequence element represented by said first icon is displayed, and

when said first icon is displayed in the sequence view, an amino acid sequence represented by said first icon is displayed and, for each respective amino acid in said amino acid sequence,

each possible codon for said respective amino acid is displayed;

the actual codon used by the open reading frame element is indicated in a first color when the actual codon has a frequency in a codon bias table that is above the codon usage threshold; and

the actual codon used by the open reading frame element is indicated in a second color when the actual codon has a frequency in the codon bias table that is below the codon usage threshold.

46. The computer program product of claim 45, wherein the amino acid sequence represented by the first icon is determined by a (i) nucleic acid sequence represented by the first icon and (ii) a reading frame of a second icon that precedes or follows said first icon in the linear or near linear arrangement, wherein said second icon represents an amino acid element.

47. The computer program product of claim 1, wherein said instructions for displaying a set of icons further comprise instructions for displaying an icon in said plurality of icons in an icon view or a sequence view, wherein,

when said icon is displayed in said icon view, a graphical depiction of the sequence element represented by said icon is displayed, and

when said icon is displayed in said sequence view, a first sequence represented by said icon is displayed, wherein said first sequence is a nucleic acid sequence or an amino acid sequence represented by said icon;

and

when said icon is displayed in said sequence view, all or a portion of the design nucleic acid sequence is displayed above said first sequence, wherein said design nucleic acid sequence corresponds to all or a portion of the amino acid sequence segments and/or the nucleic acid sequence segments of said set of sequence elements.

48. The computer program product of claim 47, wherein said first sequence is an amino acid sequence and said computer program mechanism further comprises instructions for back-translating all or a portion of said first sequence thereby constructing a corresponding portion of the design nucleic acid sequence.

49. The computer program product of claim 47, wherein said first sequence is an amino acid sequence and said computer program mechanism further comprises:

instructions for positioning a graphic icon at a position in said first sequence where a restriction site can be introduced without changing said first sequence; and

instructions for placing the restriction site recognition sequence in a corresponding position in said second design nucleic acid sequence when a user selects said graphic icon to indicate that the restriction site is desired.

50. The computer program product of claim 49, wherein said computer program mechanism further comprises:

instructions for graphically displaying overhangs generated by cleavage of the restriction site recognition sequence; and

instructions for displaying the name of the restriction enzyme that recognizes the restriction site recognition sequence in the vicinity of the restriction site recognition sequence in the design nucleic acid sequence.

51. The computer program product of claim 49, wherein,

said instructions for displaying further comprise, for each respective amino acid in said first sequence, instructions for displaying each codon corresponding to said respective amino acid sequence below said first sequence;

and

said instructions for placing further comprise instructions for highlighting each codon below said first sequence that is in the restriction site recognition sequence when a user selects said graphic icon to indicate that the restriction site is desired.

52. The computer program product of claim 47, the computer program mechanism further comprising:

a T_m calculation module, the T_m calculation module comprising;

instructions for selecting a start point and an end point in said design nucleic acid sequence, thereby defining an oligonucleotide;

instructions for computing a T_m of the oligonucleotide; and

instructions for displaying the T_m of the oligonucleotide.

53. The computer program product of claim 52, wherein the instructions for displaying the T_m of the oligonucleotide comprise instructions for displaying the T_m and a numeric representation of the start point and the end point.

54. The computer program product of claim 52, wherein the T_m calculation module further comprises instructions for moving the start point and/or the end point and, for each new start point and/or end point specified by the user, repeating said instructions for computing and said instructions for displaying.

55. The computer program product of claim 47, the computer program mechanism further comprising:

an oligonucleotide marker module, the oligonucleotide marker module comprising;

instructions for selecting a start point and an end point in said design nucleic acid sequence, thereby defining an oligonucleotide;

instructions for defining a 5′

to 3′

direction of the oligonucleotide; and

instructions for displaying the oligonucleotide as a graphic above or below the design nucleic acid sequence.

56. The computer program product of claim 1, wherein said instructions for displaying a set of icons further comprise instructions for displaying a plurality of icons in an icon view or a sequence view, wherein,

when said plurality of icons are displayed in said icon view, a graphical depiction of the set of sequence elements represented by said plurality of icons is displayed, and

when said plurality of icons are displayed in said sequence view, a plurality of sequences, each represented by one of said plurality of icons is displayed, wherein each sequence is a nucleic acid sequence or an amino acid sequence;

and

when said plurality of icons are displayed in sequence view, said design nucleic acid sequence is displayed above said plurality of sequences.

57. The computer program product of claim 1, the computer program mechanism further comprising:

instructions for merging a first sequence element and a second sequence element in said set of sequence elements thereby forming a single sequence element in said set of sequence elements from said first sequence element and said second sequence element.

58. The computer program product of claim 1, the computer program mechanism further comprising:

instructions for selecting a portion of a first sequence element in said set of sequence elements and splitting said portion of said first sequence element into a new second sequence element in said set of sequence elements.

59. The computer program product of claim 1, the computer program mechanism further comprising:

instructions for selecting a contiguous sequence wherein said contiguous sequence is all or a portion of two or more adjacent sequence elements in said linear or near linear arrangement; and

instructions for splitting said contiguous sequence into a new sequence element in said set of sequence elements and eliminating said contiguous sequence in said two or more adjacent sequence elements.

60. The computer program product of claim 1, the computer program mechanism further comprising:

instructions for saving said set of sequence elements as a first project;

instructions for permitting the selection of a project from among a plurality of projects;

each project in the plurality of projects comprising a set of sequence elements; and

instructions for linking a first sequence element in the set of sequence elements in the first project with a corresponding second sequence element in a set of sequence elements in another project in said plurality of projects such that, when changes are made to a nucleic acid sequence associated with said first sequence element, the same changes are made to a nucleic acid sequence associated with the second sequence element.

61. The computer program product of claim 60, the computer program mechanism further comprising:

instructions for removing the link between the first sequence element and the second sequence element.

62. The computer program product of claim 60, the computer program mechanism further comprising:

instructions for locking the nucleic acid sequence associated with said first sequence element and the nucleic acid sequence associated with said second sequence element so that no change is allowed to either nucleic acid sequence.

63. The computer program product of claim 1, the computer program mechanism further comprising:

instructions for altering a directional property of a sequence element in said set of sequence elements.

64. The computer program product of claim 63, wherein the directional property is a translation direction or a transcription direction.

65. The computer program product of claim 1, the computer program product further comprising:

instructions for generating a report, the report comprising any combination of;

(i) the design nucleic acid sequence;

(ii) a nucleic acid sequence associated with each sequence element in said set of sequence elements;

(iii) a codon translation map for the design nucleic acid sequence;

(iv) a restriction site summary for the design nucleic acid sequence;

(v) a codon usage frequency analysis for the design nucleic acid sequence;

(vi) a GC content for the design nucleic acid sequence;

(vii) a list of repeats in the design nucleic acid sequence; and

(viii) a list of each oligonucleotide associated with the set of sequence elements.

66. The computer program product of claim 1, wherein a first sequence element in said set of sequence elements is a DNA element and a second sequence element in said set of sequence elements in a first amino acid element, wherein said instructions for displaying a set of icons further comprise instructions for displaying a first icon in said plurality of icons representing said DNA element in an icon view or a sequence view, wherein,

when said first icon is displayed in the icon view, a graphical depiction of the sequence element represented by said first icon is displayed, and

when said first icon is displayed in the sequence view, all six reading frames of said DNA element is displayed, and wherein a first reading frame in the six reading frames that is in frame with the first amino acid element is highlighted in a first manner.

67. The computer program product of claim 66, wherein a third sequence element in said set of sequence elements is an amino acid element, and wherein when said first icon is displayed in the sequence view, a second frame in the six reading frames that is in frame with the second amino acid element is highlighted in a second manner.

68. The computer program product of claim 1, the computer program product further comprising:

instructions for communicating a sequence of the design nucleic acid sequence across a network as part of an order for said design nucleic acid sequence.

69. The computer program product of claim 68, wherein the network in the Internet.

70. The computer program product of claim 68, wherein the sequence is encrypted.

71. A computer system comprising:

a central processing unit;

a memory, coupled to the central processing unit, the memory storing;

instructions for representing a set of sequence elements, each sequence element representing an amino acid sequence segment or a nucleic acid sequence segment, wherein the set of sequence elements are collectively represented by a design nucleic acid sequence, wherein

said instructions for representing said set of sequence elements comprise instructions for displaying a plurality of icons in a linear or a near linear arrangement, each respective icon in said plurality of icons uniquely representing a corresponding sequence element in said set of sequence elements such that neighboring icons in said plurality of icons represent neighboring sequence elements in said plurality of sequence elements, and

each said respective icon in said plurality of icons depicts a directional property for the corresponding sequence element in said set of sequence elements.