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公开(公告)号：US08361720B2
公开(公告)日：2013-01-29
申请号：US12946737
申请日：2010-11-15
CPC分类号： C12Q1/6827 , C12Q1/6818 , C12Q1/686 , C12Q1/6886 , C12Q2600/156 , C12Q2600/158 , C12Q2525/301 , C12Q2527/101 , C12Q2561/109 , C12Q2565/1015
摘要： A cleavage-based real-time PCR assay method is provided. In general terms, the assay method includes subjecting a reaction mixture comprising a) PCR reagents for amplifying a nucleic acid target, and b) flap cleavage reagents for performing a flap cleavage assay on the amplified nucleic acid target to two sets of thermocycling conditions. No additional reagents are added to the reaction between said first and second sets of cycles and, in each cycle of the second set of cycles, cleavage of a flap probe is measured.
摘要翻译： 提供基于切割的实时PCR测定方法。 一般而言，测定方法包括对包含a）PCR试剂的反应混合物进行核酸靶扩增，以及b）用于在扩增的核酸靶上进行皮瓣裂解测定的折叠裂解试剂至两组热循环条件。 在所述第一组和第二组循环之间没有向所述反应中加入另外的试剂，并且在第二组循环的每个循环中，测量瓣探针的切割。
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公开(公告)号：US20130231256A1
公开(公告)日：2013-09-05
申请号：US13594674
申请日：2012-08-24
IPC分类号： C12Q1/68
CPC分类号： C12Q1/6886 , C12Q1/6858 , C12Q2600/156 , C12Q2600/16 , C12Q2525/161 , C12Q2561/109 , C12Q2565/1015
摘要： Provided herein is reagent mixture comprising multiplexed amplification reagents and flap assay reagents for detecting, in a single reaction, mutant copies of the KRAS gene that contain any of the 34A, 34C, 34T, 35A, 35C, 35T or 38A point mutations. Methods that employ the reagent mix and kits for performing the same are also provided.
摘要翻译： 本文提供的试剂混合物包含多重扩增试剂和皮瓣测定试剂，用于在单次反应中检测含有34A，34C，34T，35A，35C，35T或38A点突变中任何一种的KRAS基因的突变体拷贝。 还提供了使用试剂混合物和试剂盒进行其的方法。
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公开(公告)号：US09127318B2
公开(公告)日：2015-09-08
申请号：US13594674
申请日：2012-08-24
IPC分类号： C12Q1/68
CPC分类号： C12Q1/6886 , C12Q1/6858 , C12Q2600/156 , C12Q2600/16 , C12Q2525/161 , C12Q2561/109 , C12Q2565/1015
摘要： Provided herein is reagent mixture comprising multiplexed amplification reagents and flap assay reagents for detecting, in a single reaction, mutant copies of the KRAS gene that contain any of the 34A, 34C, 34T, 35A, 35C, 35T or 38A point mutations. Methods that employ the reagent mix and kits for performing the same are also provided.
摘要翻译： 本文提供的试剂混合物包含多重扩增试剂和皮瓣测定试剂，用于在单次反应中检测含有34A，34C，34T，35A，35C，35T或38A点突变中任何一种的KRAS基因的突变体拷贝。 还提供了使用试剂混合物和试剂盒进行其的方法。
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公开(公告)号：US20120122105A1
公开(公告)日：2012-05-17
申请号：US12946737
申请日：2010-11-15
CPC分类号： C12Q1/6827 , C12Q1/6818 , C12Q1/686 , C12Q1/6886 , C12Q2600/156 , C12Q2600/158 , C12Q2525/301 , C12Q2527/101 , C12Q2561/109 , C12Q2565/1015
摘要： A cleavage-based real-time PCR assay method is provided. In general terms, the assay method includes subjecting a reaction mixture comprising a) PCR reagents for amplifying a nucleic acid target, and b) flap cleavage reagents for performing a flap cleavage assay on the amplified nucleic acid target to two sets of thermocycling conditions. No additional reagents are added to the reaction between said first and second sets of cycles and, in each cycle of the second set of cycles, cleavage of a flap probe is measured.
摘要翻译： 提供基于切割的实时PCR测定方法。 一般而言，测定方法包括对包含a）PCR试剂的反应混合物进行核酸靶扩增，以及b）用于在扩增的核酸靶上进行皮瓣切割测定的折叠裂解试剂至两组热循环条件。 在所述第一组和第二组循环之间没有向所述反应中加入另外的试剂，并且在第二组循环的每个循环中，测量瓣探针的切割。
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公开(公告)号：US08916344B2
公开(公告)日：2014-12-23
申请号：US12946745
申请日：2010-11-15
IPC分类号： C12Q1/68
CPC分类号： C12Q1/6886 , C12Q1/6827 , C12Q2600/154 , C12Q2521/301 , C12Q2523/125 , C12Q2561/109
摘要： A method for detecting a methylated genomic locus is provided. In certain embodiments, the method comprises: a) treating a nucleic acid sample that contains both unmethylated and methylated copies of a genomic locus with an agent that modifies cytosine to uracil to produce a treated nucleic acid; b) amplifying a product from the treated nucleic acid using a first primer and a second primer, wherein the first primer hybridizes to a site in the locus that contain methylcytosines and the amplifying preferentially amplifies the methylated copies of the genomic locus, to produce an amplified sample; and c) detecting the presence of amplified methylated copies of the genomic locus in the amplified sample using a flap assay that employs an invasive oligonucleotide having a 3′ terminal G or C nucleotide that corresponds to a site of methylation in the genomic locus.
摘要翻译： 提供了一种检测甲基化基因组座位的方法。 在某些实施方案中，所述方法包括：a）将含有基因组座位的非甲基化和甲基化拷贝的核酸样品用修饰胞嘧啶至尿嘧啶的试剂处理以产生经处理的核酸; b）使用第一引物和第二引物扩增来自经处理的核酸的产物，其中第一引物与含有甲基胞嘧啶的位点的位点杂交，并且扩增优先扩增基因组基因座的甲基化拷贝，以产生扩增 样品; 和c）使用采用具有对应于基因组基因座中的甲基化位点的3'末端G或C核苷酸的侵入性寡核苷酸的皮瓣测定来检测扩增样品中扩增的基因组座位的扩增的甲基化拷贝的存在。
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公开(公告)号：US20120122106A1
公开(公告)日：2012-05-17
申请号：US12946752
申请日：2010-11-15
IPC分类号： C12Q1/68
CPC分类号： C12Q1/6886 , C12Q1/6858 , C12Q1/6881 , C12Q2600/156 , C12Q2600/158 , C12Q2525/301 , C12Q2561/109 , C12Q2565/1015
摘要： A method of sample analysis is provided. In certain embodiments, the method involves: a) amplifying a product from a sample that comprises both wild type copies of a genomic locus and mutant copies of the genomic locus that have a point mutation relative to said wild type copies of the genomic locus, to produce an amplified sample, where: i. the amplifying is done using a first primer and a second primer; and ii. the first primer comprises a 3′ terminal nucleotide that base pairs with the point mutation and also comprises a nucleotide sequence that is fully complementary to a sequence in the locus with the exception of a single base mismatch within 6 bases of the 3′ terminal nucleotide; and b) detecting the presence of said product in said amplified sample using a flap assay that employs an invasive oligonucleotide. A kit for performing the method is also provided.
摘要翻译： 提供了样品分析的方法。 在某些实施方案中，该方法包括：a）从包含基因组基因组的野生型拷贝和相对于所述基因组基因座的所述野生型拷贝的点突变的基因组基因座的突变拷贝的样品扩增产物， 产生扩增样品，其中：i。 使用第一引物和第二引物进行扩增; 和ii。 第一引物包含与点突变碱基对的3'末端核苷酸，并且还包含与3'末端核苷酸的6个碱基内的单碱基错配除外的基因座中的序列完全互补的核苷酸序列; 和b）使用采用侵入性寡核苷酸的皮瓣测定来检测所述扩增样品中所述产物的存在。 还提供了用于执行该方法的套件。
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公开(公告)号：US20120122088A1
公开(公告)日：2012-05-17
申请号：US12946745
申请日：2010-11-15
IPC分类号： C12Q1/68
CPC分类号： C12Q1/6886 , C12Q1/6827 , C12Q2600/154 , C12Q2521/301 , C12Q2523/125 , C12Q2561/109
摘要： A method for detecting a methylated genomic locus is provided. In certain embodiments, the method comprises: a) treating a nucleic acid sample that contains both unmethylated and methylated copies of a genomic locus with an agent that modifies cytosine to uracil to produce a treated nucleic acid; b) amplifying a product from the treated nucleic acid using a first primer and a second primer, wherein the first primer hybridizes to a site in the locus that contain methylcytosines and the amplifying preferentially amplifies the methylated copies of the genomic locus, to produce an amplified sample; and c) detecting the presence of amplified methylated copies of the genomic locus in the amplified sample using a flap assay that employs an invasive oligonucleotide having a 3′ terminal G or C nucleotide that corresponds to a site of methylation in the genomic locus.
摘要翻译： 提供了一种检测甲基化基因组座位的方法。 在某些实施方案中，所述方法包括：a）将含有基因组座位的非甲基化和甲基化拷贝的核酸样品用修饰胞嘧啶至尿嘧啶的试剂处理以产生经处理的核酸; b）使用第一引物和第二引物扩增来自经处理的核酸的产物，其中第一引物与含有甲基胞嘧啶的位点的位点杂交，并且扩增优先扩增基因组基因座的甲基化拷贝，以产生扩增的 样品; 和c）使用采用具有对应于基因组基因座中的甲基化位点的3'末端G或C核苷酸的侵入性寡核苷酸的皮瓣测定来检测扩增样品中扩增的基因组座位的扩增的甲基化拷贝的存在。
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公开(公告)号：US08715937B2
公开(公告)日：2014-05-06
申请号：US12946752
申请日：2010-11-15
CPC分类号： C12Q1/6886 , C12Q1/6858 , C12Q1/6881 , C12Q2600/156 , C12Q2600/158 , C12Q2525/301 , C12Q2561/109 , C12Q2565/1015
摘要： A method of sample analysis is provided. In certain embodiments, the method involves: a) amplifying a product from a sample that comprises both wild type copies of a genomic locus and mutant copies of the genomic locus that have a point mutation relative to the wild type copies of the genomic locus, to produce an amplified sample, where: i. the amplifying is done using a first primer and a second primer; and ii. the first primer comprises a 3′ terminal nucleotide that base pairs with the point mutation and also comprises a nucleotide sequence that is fully complementary to a sequence in the locus with the exception of a single base mismatch within 6 bases of the 3′ terminal nucleotide; and b) detecting the presence of the product in the amplified sample using a flap assay that employs an invasive oligonucleotide. A kit for performing the method is also provided.
摘要翻译： 提供了样品分析的方法。 在某些实施方案中，所述方法包括：a）从包含基因组基因组的野生型拷贝和相对于基因组基因座的野生型拷贝的点突变的基因组基因座的突变拷贝的样品扩增产物， 产生扩增样品，其中：i。 使用第一引物和第二引物进行扩增; 和ii。 第一引物包含与点突变碱基对的3'末端核苷酸，并且还包含与3'末端核苷酸的6个碱基内的单碱基错配除外的基因座中的序列完全互补的核苷酸序列; 和b）使用采用侵入性寡核苷酸的皮瓣测定法检测扩增样品中产物的存在。 还提供了用于执行该方法的套件。
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公开(公告)号：US20130196322A1
公开(公告)日：2013-08-01
申请号：US13754631
申请日：2013-01-30
申请人： Michael J. Domanico , Hatim Allawi , Graham P. Lidgard , Brian Aizenstein , Oliver Hunt , Tobias Charles Zutz
发明人： Michael J. Domanico , Hatim Allawi , Graham P. Lidgard , Brian Aizenstein , Oliver Hunt , Tobias Charles Zutz
IPC分类号： C12Q1/68
CPC分类号： C12Q1/6806 , C12Q1/6851 , C12Q2600/154 , C12Q2523/113 , C12Q2523/125 , C12Q2527/125 , C12Q2527/137
摘要： Provided herein is technology related to the chemical modification and purification of DNA. Specifically, the technology provides methods for performing a bisulfate conversion reaction on small amounts of single-stranded, fragmented DNA and performing the subsequent desulfonation and purification steps on magnetic beads.
摘要翻译： 本文提供了与DNA的化学修饰和纯化有关的技术。 具体地，该技术提供了对少量单链，片段化的DNA进行硫酸氢盐转化反应并在磁珠上进行随后的脱磺化和纯化步骤的方法。
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公开(公告)号：US20120288868A1
公开(公告)日：2012-11-15
申请号：US13470251
申请日：2012-05-11
申请人： Janelle J. Bruinsma , Michael J. Domanico , Graham P. Lidgard , Hongzhi Zou , William G. Weisburg , Hemanth D. Shenoi , James P. Light, II
发明人： Janelle J. Bruinsma , Michael J. Domanico , Graham P. Lidgard , Hongzhi Zou , William G. Weisburg , Hemanth D. Shenoi , James P. Light, II
CPC分类号： C12Q1/6806 , B01D33/15 , B01D33/155 , B01L3/00 , B01L3/5021 , B03C1/30 , B04B3/00 , C12N15/1006 , C12N15/1013 , C12N15/1017 , C12Q1/6886 , C12Q2600/158 , C12Q2600/16 , Y10T436/143333
摘要： Provided herein is technology relating to isolating nucleic acids. In particular, the technology relates to methods and kits for extracting nucleic acids from problematic samples such as stool.
摘要翻译： 本文提供了与分离核酸有关的技术。 特别地，该技术涉及从有问题的样品如粪便中提取核酸的方法和试剂盒。
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