公开(公告)号：US09765385B2

公开(公告)日：2017-09-19

申请号：US14411806

申请日：2013-06-28

Applicant: HTG Molecular Diagnostics, Inc.

Inventor： Matt Rounseville ,  Bruce Seligmann

IPC: C12Q1/68

CPC classification number: C12Q1/683 ,  C12Q1/6827 ,  C12Q2521/325 ,  C12Q2561/108 ,  C12Q2537/143 ,  C12Q2563/107

Abstract: Disclosed herein are methods for detecting presence of a nucleotide variant in a target nucleic acid utilizing a nuclease protection assay. The methods include contacting a sample with at least two probes, wherein the first probe is complementary to the wild-type (non-variant) nucleotide(s) at the nucleotide variant position(s) in the target nucleic acid and the second probe is complementary to the variant nucleotide(s) at the nucleotide variant position(s) in the target nucleic acid, under conditions sufficient for the probes to hybridize to the target nucleic acid, producing a mixture of hybridized and unhybridized nucleic acids. The mixture is contacted with a nuclease specific for single-stranded nucleic acid molecules under conditions sufficient to remove unhybridized nucleic acid molecules (or unhybridized portions of nucleic acid molecules). The presence of the at least two probes is then detected, thereby detecting the presence of the variant and/or non-variant target nucleic acid in the sample.

公开(公告)号：US20150191770A1

公开(公告)日：2015-07-09

申请号：US14411806

申请日：2013-06-28

Applicant: HTG Molecular Diagnostics, Inc.

Inventor： Matt Rounseville ,  Bruce Seligmann

IPC: C12Q1/68

CPC classification number: C12Q1/683 ,  C12Q1/6827 ,  C12Q2521/325 ,  C12Q2561/108 ,  C12Q2537/143 ,  C12Q2563/107

Abstract: Disclosed herein are methods for detecting presence of a nucleotide variant in a target nucleic acid utilizing a nuclease protection assay. The methods include contacting a sample with at least two probes, wherein the first probe is complementary to the wild-type (non-variant) nucleotide(s) at the nucleotide variant position(s) in the target nucleic acid and the second probe is complementary to the variant nucleotide(s) at the nucleotide variant position(s) in the target nucleic acid, under conditions sufficient for the probes to hybridize to the target nucleic acid, producing a mixture of hybridized and unhybridized nucleic acids. The mixture is contacted with a nuclease specific for single-stranded nucleic acid molecules under conditions sufficient to remove unhybridized nucleic acid molecules (or unhybridized portions of nucleic acid molecules). The presence of the at least two probes is then detected, thereby detecting the presence of the variant and/or non-variant target nucleic acid in the sample.

Abstract translation: 本文公开了使用核酸酶保护测定法检测靶核酸中核苷酸变体的存在的方法。 所述方法包括使样品与至少两个探针接触，其中第一探针与靶核酸中核苷酸变体位置处的野生型（非变体）核苷酸互补，第二探针是 与靶核酸的核苷酸变体位置的变体核苷酸互补，在足以使探针与靶核酸杂交的条件下，产生杂交和未杂交的核酸的混合物。 在足以除去未杂交的核酸分子（或核酸分子的未杂交部分）的条件下，将混合物与单链核酸分子特异性核酸酶接触。 然后检测至少两个探针的存在，从而检测样品中变体和/或非变体靶核酸的存在。

公开(公告)号：US09512469B2

公开(公告)日：2016-12-06

申请号：US14347855

申请日：2012-09-26

Applicant: HTG Molecular Diagnostics, Inc.

Inventor： Bruce Seligmann ,  Matt Rounseville ,  Krishna Maddula ,  Ihab Botros ,  Chris Cox

IPC: C12Q1/68 ,  C12M1/34 ,  C12N9/22 ,  C07H21/02 ,  C07H21/04 ,  G01N31/22 ,  C40B30/04

CPC classification number: C12Q1/6816 ,  C12Q2521/325 ,  C12Q2525/207 ,  C12Q2537/143 ,  C12Q2561/108

Abstract: Disclosed herein are methods of co-detecting presence of target messenger RNA (mRNA) and small non-coding RNA (for example, miRNA) in a sample. The disclosed methods can be used to simultaneously detect mRNA and small non-coding RNA in a single assay (for example in the same reaction or the same well of a multi-well assay). The methods can include contacting a sample with a plurality of nuclease protection probes (NPPs) including at least one probe which specifically binds to a target mRNA and at least one probe which specifically binds to a target small non-coding RNA, contacting the sample with a nuclease specific for single-stranded nucleic acids, and detecting the NPP, for example on a microarray.

Abstract translation: 本文公开了共检测样品中靶信使RNA（mRNA）和小非编码RNA（例如miRNA）的存在的方法。 所公开的方法可以用于在单个测定中（例如在多孔测定的相同反应或相同的孔中）同时检测mRNA和小的非编码RNA。 所述方法可以包括使样品与多个核酸酶保护探针（NPP）接触，所述核酸酶保护探针（NPP）包括至少一个特异性结合靶mRNA的探针和至少一个特异性结合目标小非编码RNA的探针，使样品与 对单链核酸特异的核酸酶，并且例如在微阵列上检测NPP。