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    SINGLE NUCLEOTIDE ANALYTICAL METHOD AND ASSOCIATED PROBES
    1.
    发明申请
    SINGLE NUCLEOTIDE ANALYTICAL METHOD AND ASSOCIATED PROBES 审中-公开

    公开(公告)号：WO2019081483A1

    公开(公告)日：2019-05-02

    申请号：PCT/EP2018/078991

    申请日：2018-10-23

    申请人： BASE4 INNOVATION LIMITED

    发明人： BALMFORTH, Barnaby ,  FRAYLING, Cameron Alexander

    IPC分类号： C12Q1/6823

    CPC分类号： C12Q1/6823 ,  C12Q2521/101 ,  C12Q2521/301 ,  C12Q2521/319 ,  C12Q2525/117 ,  C12Q2525/125 ,  C12Q2525/307 ,  C12Q2563/159 ,  C12Q2537/149 ,  C12Q2563/107 ,  C12Q2537/164

    摘要： A method of detecting whether the nucleobase of a given nucleoside triphosphate molecule in an analyte does or does not include a given chemical or structural modification characterised by the steps of (1) reacting the analyte in the presence of a polymerase with a biological probe comprised of (a) a pair of single-stranded first oligonucleotides each comprising an exonuclease blocking-site; at least one restriction endonuclease recognition-site located on the 3' side of the 10 blocking-site; a single nucleotide capture-site located within the recognition-site and respectively first and second fluorophore(s) located on the 3' side of the recognition-site arranged so as to be substantially undetectable and (b) at least one second and optionally at least one third single- stranded oligonucleotide each separate from the first oligonucleotide and capable of hybridising to complementary flanking regions on the 3' and 5' sides of the capture-site of one, other or both 15 of the first oligonucleotides in the pair to create a used-probe duplex consisting of (b1) one or other of the first oligonucleotides in the pair and (b2) a component comprised of the second oligonucleotide, one or other of a modified or unmodified nucleotide derived from the nucleoside triphosphate molecule and optionally the third oligonucleotide wherein each duplex is comprised of one of the four possible (b1)/(b2) duplex permutations; (2) reacting the duplex produced in 20 step (1) with a restriction endonuclease system comprised of at least one restriction endonuclease adapted to selectively cleave the (b1) strand at the recognition-site to create depending on the presence or absence of the modification in the original nucleoside triphosphate molecule (i) only exonucleolytically-digestible first oligonucleotide elements bearing first fluorophore(s) or (ii) only exonucleolytically-digestible first oligonucleotide elements bearing 25 second fluorophore(s) or (iii) a mixture of exonucleolytically-digestible first oligonucleotide elements respectively bearing first and second fluorophore(s); (3) creating another used-probe duplex from the (b2) component and another first oligonucleotide in the pair and thereafter iterating steps (2) and (3); (4) digesting the first oligonucleotide elements with an enzyme having 5'-3' exonucleolytic activity to produce fluorophore(s) in a detectable state after either or both of 30 steps (2) and (3); and (5) detecting the fluorophore(s) released in step (4) and inferring from the nature of the fluorescence signal observed whether the original single nucleoside triphosphate molecule was modified or unmodified.

    MICROARRAY SYSTEM WITH IMPROVED SEQUENCE SPECIFICITY
    6.
    发明申请
    MICROARRAY SYSTEM WITH IMPROVED SEQUENCE SPECIFICITY 审中-公开
    具有改进的序列特异性的微阵列系统

    公开(公告)号：WO2009023676A1

    公开(公告)日：2009-02-19

    申请号：PCT/US2008/072924

    申请日：2008-08-12

    申请人： INTEGRATED DNA TECHNOLOGIES, INC. ,  GUNNING, Kerry, B. ,  BEHLKE, Mark, Aaron

    发明人： GUNNING, Kerry, B. ,  BEHLKE, Mark, Aaron

    IPC分类号： C12P19/34 ,  C07H21/04

    CPC分类号： C12Q1/6837 ,  B01J19/0046 ,  B01J2219/0059 ,  B01J2219/00608 ,  B01J2219/00626 ,  B01J2219/00722 ,  C12P19/34 ,  C12Q1/6806 ,  Y10T436/108331 ,  C12Q2525/125 ,  C12Q2533/101 ,  C12Q2521/319

    摘要： The invention provides a novel array method for nucleic acid sequence detection with improved specificity which allows for detection of genetic variation, from simple SNPs (where the variation occurs at a fixed position and is of limited allelic number) to more complex sequence variation patterns (such as with multigene families or multiple genetic strains of an organism where the sequence variation between the individual members is neither fixed nor consistent). The array is comprised of short, synthetic oligonucleotide probes attached to a solid surface which are hybridized to single-stranded targets. Single stranded targets can be produced using a method that employs primers modified on the 5 ' end to prohibit degradation by a 5'-exonuclease that is introduced to degrade the unprotected strand. The invention further provides for printing buffers/solutions for the immobilization of oligonucleotide probes to an array surface. The invention also provides hybridization and wash buffers and conditions to maximize hybridization specificity and signal intensity, and reduce hybridization times.

    摘要翻译： 本发明提供了一种用于核酸序列检测的新型阵列方法，其具有改进的特异性，其允许从简单SNP（其中在固定位置发生并且具有有限等位基数的变体）检测遗传变异与更复杂的序列变异模式（例如 与多基因家族或生物体的多种遗传菌株一样，其中各个成员之间的序列变异既不固定也不一致）。 阵列由连接到固体表面的短合成寡核苷酸探针组成，其与单链靶标杂交。 可以使用使用在5'端修饰的引物来防止引入降解未保护链的5'-外切核酸酶降解的方法来产生单链靶标。 本发明还提供用于将寡核苷酸探针固定到阵列表面的印刷缓冲液/溶液。 本发明还提供杂交和洗涤缓冲液和条件以使杂交特异性和信号强度最大化，并减少杂交时间。

    METHODS OF SCREENING NUCLEIC ACIDS FOR SINGLE NUCLEOTIDE VARIATIONS
    7.
    发明申请
    METHODS OF SCREENING NUCLEIC ACIDS FOR SINGLE NUCLEOTIDE VARIATIONS 审中-公开
    筛选单核细胞核变种的方法

    公开(公告)号：WO2008143903A3

    公开(公告)日：2009-01-08

    申请号：PCT/US2008006191

    申请日：2008-05-14

    申请人： INSIGHT GENETICS INC ,  DAHLHAUSER ERIC B

    发明人： DAHLHAUSER ERIC B

    IPC分类号： C12Q1/68

    CPC分类号： C12Q1/6827 ,  C12Q1/6834 ,  C12Q2535/125 ,  C12Q2525/125 ,  C12Q2521/319 ,  C12Q2521/101

    摘要： Disclosed are methods and compositions for detecting variation in nucleic acids. The disclosed method compares the sequence of a nucleic acid of interest with the sequence of a reference nucleic acid to sensitively identify variations between the sequence of a nucleic acid of interest and the sequence of a reference nucleic acid. The disclosed method generally involves excision and replacement of selected nucleotides in nucleic acid strands hybridized to other strands. In the method, if the excised nucleotide was mismatched with the nucleotide in the other, hybridized strand, then the replacement nucleotide will not be mismatched. If the excised nucleotide was not mismatched with the nucleotide in the other, hybridized strand, then the excised nucleotide is not replaced. This difference allows detection of variation in the nucleic acid of interest. In some forms of the method, by replacing excised nucleotides with nuclease-resistant nucleotides, strands in which excised nucleotides are replaced will be resistant to nuclease digestion while strands in which excised nucleotides are not replaced will be sensitive to nuclease digestion. By exposing the hybridizing nucleic acids to nuclease following replacement of excised nucleotides, the strands in which excised nucleotides are not replaced can be destroyed by the nuclease while strands in which excised nucleotides are replaced can be preserved. The remaining strands can then be detected and whether the strand survived nuclease digestion can be noted. Strands that survive nuclease digestion are indicative of the presence of variation in the nucleic acid of interest.

    摘要翻译： 公开了用于检测核酸变异的方法和组合物。 所公开的方法将感兴趣的核酸序列与参考核酸的序列进行比较以敏感地鉴定目的核酸序列与参照核酸的序列之间的变化。 所公开的方法通常涉及与其它链杂交的核酸链中的选择的核苷酸的切除和置换。 在该方法中，如果切除的核苷酸与另一个的核苷酸错配，则杂交的链，则替换核苷酸将不匹配。 如果切除的核苷酸与另一个杂交的核苷酸不匹配，那么切除的核苷酸不被替换。 该差异允许检测感兴趣的核酸中的变异。 在该方法的一些形式中，通过用核酸酶抗性核苷酸替代切除的核苷酸，其中切割的核苷酸被替换的链将对核酸酶消化具有抗性，而未被替换的切除的核苷酸的链对核酸酶消化敏感。 通过在切割的核苷酸替代后将杂交核酸暴露于核酸酶，切割的核苷酸未被替换的链可以被核酸酶破坏，而可以保留切割的核苷酸被替换的链。 然后可以检测剩余的链，并且可以注意链中是否存活核酸酶消化。 在核酸酶消化中存活的链表示感兴趣的核酸中存在变异。

    DETECTION OF TARGET NUCLEOTIDE SEQUENCES USING AN ASYMMETRIC OLIGONUCLEOTIDE LIGATION ASSAY
    8.
    发明申请
    DETECTION OF TARGET NUCLEOTIDE SEQUENCES USING AN ASYMMETRIC OLIGONUCLEOTIDE LIGATION ASSAY 审中-公开
    使用不对称寡核苷酸引物测定法检测目标核苷酸序列

    公开(公告)号：WO2005118847A1

    公开(公告)日：2005-12-15

    申请号：PCT/EP2005/005859

    申请日：2005-05-30

    申请人： KEYGENE N.V. ,  HOGERS, René, Cornelis, Josephus

    发明人： HOGERS, René, Cornelis, Josephus

    IPC分类号： C12Q1/68

    CPC分类号： C12Q1/6827 ,  C12Q1/6858 ,  C12Q1/6862 ,  C12Q2563/179 ,  C12Q2525/155 ,  C12Q2525/125 ,  C12Q2521/501 ,  C12Q2521/319 ,  C12Q2525/186 ,  C12Q2521/325

    摘要： The invention relates to a method for the detection of target nucleic acid sequences based on ligation of a first and a second probe that are hybridized adjacent on a target nucleic acid sequence, removal of any unligated probes, preferably by exonuclease treatment, amplification of a portion of the second probe that does not contain the target specific section to provide amplicons and detection of said amplicons. The second probe comprises a second target binding section, one or two primer binding sequences and an identifier. The first probe preferably does not comprise a primer binding sequence, preferably consists of a first target binding section and is preferably exonuclease resistant.

    摘要翻译： 本发明涉及一种基于在靶核酸序列上与邻近杂交的第一和第二探针的连接来检测靶核酸序列的方法，优选通过外切核酸酶处理去除任何未标记的探针，扩增部分 的第二探针，其不包含靶特异性区段以提供扩增子和所述扩增子的检测。 第二探针包括第二靶结合部分，一个或两个引物结合序列和标识符。 第一探针优选不包含引物结合序列，优选由第一靶结合部分组成并且优选为外切核酸酶抗性。

    ALLELE SPECIFIC PRIMER EXTENSION
    9.
    发明申请
    ALLELE SPECIFIC PRIMER EXTENSION 审中-公开
    ALLELE特定的PRIMER扩展

    公开(公告)号：WO2004020604A3

    公开(公告)日：2004-08-12

    申请号：PCT/US0327286

    申请日：2003-08-29

    申请人： AMERSHAM BIOSCIENCES CORP

    发明人： SOOD ANUP ,  KUMAR SHIV ,  FULLER CARL ,  NELSON JOHN ,  MACKLIN JOHN

    IPC分类号： C12N15/09 ,  C12Q1/42 ,  C12Q1/44 ,  C12Q1/68 ,  C12P19/34

    CPC分类号： C12Q1/6869 ,  C12Q1/6823 ,  C12Q2600/156 ,  C12Q2535/125 ,  C12Q2525/125 ,  C12Q2521/125 ,  C12Q2565/301 ,  C12Q2525/101

    摘要： A method of characterizing a nucleic acid sample is provided that includes the steps of: (a) conducting a DNA polymerase reaction that includes the reaction of a template, an allele specific primer, at least one terminal phosphate-labeled nucleotide, DNA polymerase, and optionally an enzyme having 3' ? 5' exonuclease activity when the primer is non-hydrolyzable, which reaction results in the production of labeled polyphosphate; (b) permitting the labeled polyphosphate to react with a phosphatase to produce a detectable species; (c) detecting the detectable species; and (d) characterizing the nucleic acid sample based on such detection.

    摘要翻译： 提供了表征核酸样品的方法，其包括以下步骤：（a）进行DNA聚合酶反应，其包括模板，等位基因特异性引物，至少一个末端磷酸酯标记的核苷酸，DNA聚合酶和 任选地具有3' 5'核酸外切酶活性，该反应导致标记多磷酸盐的产生; （b）允许标记的多磷酸酯与磷酸酶反应以产生可检测的物质; （c）检测可检测物种; 和（d）基于这种检测来表征核酸样品。

    METHOD FOR DETERMINING POLYNUCLEOTIDE SEQUENCE VARIATIONS
    10.
    发明申请
    METHOD FOR DETERMINING POLYNUCLEOTIDE SEQUENCE VARIATIONS 审中-公开
    确定多核苷酸序列变异的方法

    公开(公告)号：WO2004065634A1

    公开(公告)日：2004-08-05

    申请号：PCT/US2003/001342

    申请日：2003-01-15

    申请人： BIOVENTURES, INC. ,  DAWSON, Elliott, P.

    发明人： DAWSON, Elliott, P.

    IPC分类号： C12Q1/68

    CPC分类号： C12Q1/6858 ,  C12Q2525/125

    摘要： A method for determining the presence, location or identity, or a combination of these, of the nucleotides in a polynucleotide. A method for determining the presence, location or identity, or a combination of these, of one or more than one nucleotide difference between a first polynucleotide and a second polynucleotide, or between more than two polynucleotides.

    摘要翻译： 确定多核苷酸中核苷酸的存在，位置或同一性或这些的组合的方法。 用于确定第一多核苷酸和第二多核苷酸之间或多于两个多核苷酸之间的一个或多于一个核苷酸差异的存在，位置或同一性或这些的组合的方法。