公开(公告)号：US10954560B2

公开(公告)日：2021-03-23

申请号：US16215489

申请日：2018-12-10

Applicant: Fluidigm Corporation

Inventor： Carolyn G. Conant ,  Tze Howe Charn ,  Jason A. A. West ,  Xiaohui Wang

IPC: C12Q1/68 ,  B01L3/00 ,  C12Q1/6874 ,  C12Q1/6844

Abstract: Described herein are cell-based analytic methods, including a method of incorporating nucleic acid sequences into reaction products from a cell population, wherein the nucleic acid sequences are incorporated into the reaction products of each cell individually or in small groups of cells individually. Also described herein is a matrix-type microfluidic device that permits at least two reagents to be delivered separately to each cell or group of cells, as well as primer combinations useful in the method and device.

公开(公告)号：US20190185929A1

公开(公告)日：2019-06-20

申请号：US16215489

申请日：2018-12-10

Applicant: Fluidigm Corporation

Inventor： Carolyn G. Conant ,  Tze Howe Charn ,  Jason A. A. West ,  Xiaohui Wang

IPC: C12Q1/6874 ,  B01L3/00 ,  C12Q1/6844

CPC classification number: C12Q1/6874 ,  B01L3/5027 ,  B01L3/502761 ,  B01L2200/0652 ,  B01L2300/0609 ,  B01L2300/0893 ,  C12Q1/6844 ,  C12Q2525/179 ,  C12Q2563/179 ,  C12Q2565/629

Abstract: Described herein are cell-based analytic methods, including a method of incorporating nucleic acid sequences into reaction products from a cell population, wherein the nucleic acid sequences are incorporated into the reaction products of each cell individually or in small groups of cells individually. Also described herein is a matrix-type microfluidic device that permits at least two reagents to be delivered separately to each cell or group of cells, as well as primer combinations useful in the method and device.

公开(公告)号：US09644231B2

公开(公告)日：2017-05-09

申请号：US14713887

申请日：2015-05-15

Applicant: Fluidigm Corporation

Inventor： Kenneth J. Livak ,  Stacey N. Meyers ,  Xiaohui Wang ,  Jun Wang

IPC: G01N21/64 ,  C40B30/04 ,  C12Q1/68

CPC classification number: C12Q1/6818 ,  C12Q2525/151

Abstract: The invention provides a method for detecting a target nucleotide sequence by tagging the nucleotide sequence with a nucleotide tag, providing a probe oligonucleotide with a melting temperature Tm1, comprising a regulatory sequence and a nucleotide tag recognition sequence; incorporating the probe oligonucleotide into the tagged polynucleotide in a polynucleotide amplification reaction, providing a regulatory oligonucleotide with a melting temperature Tm2, comprising a sequence segment that complementary to the regulatory sequence and a tail segment that does not hybridize to the probe nucleotide when the sequence segment and the regulatory sequence are annealed, amplifying the tagged target nucleic acid sequence in a PCR amplification reaction using the probe oligonucleotide as a primer, and using a DNA polymerase with high strand displacement activity and low 5′-nuclease activity, and detecting the amplification product; wherein Tm1 and Tm2 are higher than the annealing temperature associated with the polynucleotide amplification reaction.

公开(公告)号：US09587272B2

公开(公告)日：2017-03-07

申请号：US14340446

申请日：2014-07-24

Applicant: Fluidigm Corporation

Inventor： Kenneth J. Livak ,  Stacey N. Meyers ,  Jun Wang ,  Xiaohui Wang

IPC: C07H21/04 ,  C12Q1/68

CPC classification number: C12Q1/6818 ,  C12Q2525/151

Abstract: The invention provides a method for detecting a target nucleotide sequence by tagging the nucleotide sequence with a nucleotide tag, providing a probe oligonucleotide with a melting temperature Tm1, comprising a regulatory sequence and a nucleotide tag recognition sequence; incorporating the probe oligonucleotide into the tagged polynucleotide in a polynucleotide amplification reaction, providing a regulatory oligonucleotide with a melting temperature Tm2, comprising a sequence segment that is at least partially complementary to the regulatory sequence, amplifying the tagged target nucleic acid sequence in a PCR amplification reaction using the probe oligonucleotide as a primer, and detecting the amplification product; wherein Tm1 and Tm2 are higher than the annealing temperature associated with the polynucleotide amplification reaction.

Abstract translation: 本发明提供了通过用核苷酸标签标记核苷酸序列来检测靶核苷酸序列的方法，提供了包含调节序列和核苷酸标签识别序列的具有解链温度Tm1的探针寡核苷酸; 在多核苷酸扩增反应中将探针寡核苷酸掺入标记的多核苷酸中，提供具有解链温度Tm2的调节性寡核苷酸，其包含与调节序列至少部分互补的序列片段，在PCR扩增中扩增标记的靶核酸序列 使用探针寡核苷酸作为引物的反应，检测扩增产物; 其中Tm1和Tm2高于与多核苷酸扩增反应相关的退火温度。

公开(公告)号：US20150337298A1

公开(公告)日：2015-11-26

申请号：US14719149

申请日：2015-05-21

Applicant: FLUIDIGM CORPORATION

Inventor： Lei Xi ,  Xiaohui Wang ,  Marc Unger ,  David Ruff

IPC: C12N15/10 ,  C12Q1/68

CPC classification number: C12N15/1082 ,  C12N15/1065 ,  C12Q1/6806 ,  C12Q2525/155 ,  C12Q2525/301 ,  C12Q2535/122

Abstract: In certain embodiments, the present invention provides a way of “digitally” marking different the alleles of different chromosomes by using a transposase to insert differently barcoded transposons into genomic DNA before further analysis. According to this method, each allele becomes marked with a unique pattern of transposon barcodes. Because each unique pattern of transposon barcodes identifies a particular allele, the method facilitates determinations of ploidy and copy number variation, improves the ability to discriminate among homozygotes, heterozygotes, and patterns arising from sequencing errors, and allows loci separated by uninformative stretches of DNA to be identified as linked loci, thereby facilitating haplotype determinations. Also provided is a novel artificial transposon end that includes a barcode sequence in two or more positions that are not essential for transposition.

Abstract translation: 在某些实施方案中，本发明提供了一种通过使用转座酶在进一步分析之前将不同条形码转座子插入基因组DNA中来“不同”地标记不同染色体等位基因的方式。 根据这种方法，每个等位基因都被标记为转座子条形码的独特模式。 因为转座子条形码的每个独特模式识别特定的等位基因，所以该方法有助于确定倍性和拷贝数变异，提高了鉴别纯合子，杂合子和测序错误引起的模式的能力，并且允许通过DNA的非信息延伸分离的位点 被确定为相关基因座，从而促进单倍型测定。 还提供了一种新颖的人工转座子末端，其包括对于转座不是必需的两个或更多个位置的条形码序列。

公开(公告)号：US20150147755A1

公开(公告)日：2015-05-28

申请号：US14340446

申请日：2014-07-24

Applicant: Fluidigm Corporation

Inventor： Kenneth J. Livak ,  Stacey N. Myers ,  Jun Wang ,  Xiaohui Wang

IPC: C12Q1/68

CPC classification number: C12Q1/6818 ,  C12Q2525/151

Abstract: The invention provides a method for detecting a target nucleotide sequence by tagging the nucleotide sequence with a nucleotide tag, providing a probe oligonucleotide with a melting temperature Tm1, comprising a regulatory sequence and a nucleotide tag recognition sequence; incorporating the probe oligonucleotide into the tagged polynucleotide in a polynucleotide amplification reaction, providing a regulatory oligonucleotide with a melting temperature Tm2, comprising a sequence segment that is at least partially complementary to the regulatory sequence, amplifying the tagged target nucleic acid sequence in a PCR amplification reaction using the probe oligonucleotide as a primer, and detecting the amplification product; wherein Tm1 and Tm2 are higher than the annealing temperature associated with the polynucleotide amplification reaction.

Abstract translation: 本发明提供了通过用核苷酸标签标记核苷酸序列来检测靶核苷酸序列的方法，提供了具有调节序列和核苷酸标签识别序列的融解温度Tm1的探针寡核苷酸; 在多核苷酸扩增反应中将探针寡核苷酸掺入标记的多核苷酸中，提供具有解链温度Tm2的调节性寡核苷酸，其包含与调节序列至少部分互补的序列片段，在PCR扩增中扩增标记的靶核酸序列 使用探针寡核苷酸作为引物的反应，检测扩增产物; 其中Tm1和Tm2高于与多核苷酸扩增反应相关的退火温度。

公开(公告)号：US10190163B2

公开(公告)日：2019-01-29

申请号：US15055252

申请日：2016-02-26

Applicant: Fluidigm Corporation

Inventor： Carolyn G. Conant ,  Tze Howe Charn ,  Jason A. A. West ,  Xiaohui Wang

IPC: C12Q1/68 ,  C12Q1/6874 ,  C12Q1/6844 ,  B01L3/00

Abstract: Described herein are cell-based analytic methods, including a method of incorporating nucleic acid sequences into reaction products from a cell population, wherein the nucleic acid sequences are incorporated into the reaction products of each cell individually or in small groups of cells individually. Also described herein is a matrix-type microfluidic device that permits at least two reagents to be delivered separately to each cell or group of cells, as well as primer combinations useful in the method and device.

公开(公告)号：US20160251714A1

公开(公告)日：2016-09-01

申请号：US15055252

申请日：2016-02-26

Applicant: Fluidigm Corporation

Inventor： Carolyn G. Conant ,  Tze Howe Charn ,  Jason A. A. West ,  Xiaohui Wang

IPC: C12Q1/68

CPC classification number: C12Q1/6874 ,  B01L3/502761 ,  B01L2200/0652 ,  B01L2300/0609 ,  B01L2300/0893 ,  C12Q1/6844 ,  C12Q2525/179 ,  C12Q2563/179 ,  C12Q2565/629

Abstract: Described herein are cell-based analytic methods, including a method of incorporating nucleic acid sequences into reaction products from a cell population, wherein the nucleic acid sequences are incorporated into the reaction products of each cell individually or in small groups of cells individually. Also described herein is a matrix-type microfluidic device that permits at least two reagents to be delivered separately to each cell or group of cells, as well as primer combinations useful in the method and device.

Abstract translation: 本文描述的是基于细胞的分析方法，包括将核酸序列并入来自细胞群体的反应产物中的方法，其中所述核酸序列单独地或以小组细胞单独引入每个细胞的反应产物中。 本文还描述了允许至少两种试剂分别递送至每个细胞或细胞组的基质型微流体装置，以及在该方法和装置中有用的引物组合。

公开(公告)号：US20150361486A1

公开(公告)日：2015-12-17

申请号：US14713887

申请日：2015-05-15

Applicant: FLUIDIGM CORPORATION

Inventor： Kenneth J. Livak ,  Stacey N. Meyers ,  Xiaohui Wang ,  Jun Wang

IPC: C12Q1/68

CPC classification number: C12Q1/6818 ,  C12Q2525/151

Abstract: The invention provides a method for detecting a target nucleotide sequence by tagging the nucleotide sequence with a nucleotide tag, providing a probe oligonucleotide with a melting temperature Tm1, comprising a regulatory sequence and a nucleotide tag recognition sequence; incorporating the probe oligonucleotide into the tagged polynucleotide in a polynucleotide amplification reaction, providing a regulatory oligonucleotide with a melting temperature Tm2, comprising a sequence segment that complementary to the regulatory sequence and a tail segment that does not hybridize to the probe nucleotide when the sequence segment and the regulatory sequence are annealed, amplifying the tagged target nucleic acid sequence in a PCR amplification reaction using the probe oligonucleotide as a primer, and using a DNA polymerase with high strand displacement activity and low 5′-nuclease activity, and detecting the amplification product; wherein Tm1 and Tm2 are higher than the annealing temperature associated with the polynucleotide amplification reaction.

Abstract translation: 本发明提供了通过用核苷酸标签标记核苷酸序列来检测靶核苷酸序列的方法，提供了具有调节序列和核苷酸标签识别序列的融解温度Tm1的探针寡核苷酸; 在多核苷酸扩增反应中将探针寡核苷酸掺入标记的多核苷酸，提供具有解链温度Tm2的调节性寡核苷酸，其包含与调节序列互补的序列区段和当序列片段不与探针核苷酸杂交时的尾段 并使用探针寡核苷酸作为引物，并使用具有高链置换活性和低5'-核酸酶活性的DNA聚合酶，在PCR扩增反应中扩增标记的靶核酸序列，并检测扩增产物 ; 其中Tm1和Tm2高于与多核苷酸扩增反应相关的退火温度。