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公开(公告)号：US06569647B1
公开(公告)日：2003-05-27
申请号：US09299217
申请日：1999-04-23
IPC分类号： C12P1934
CPC分类号： C12Q1/686 , C12Q1/6816 , C12Q1/6834 , C12Q1/6862 , C12Q1/689 , Y10T436/143333 , C12Q2565/543 , C12Q2563/143 , C12Q2525/185 , C12Q2525/307 , C12Q2523/101 , C12Q2563/131 , C12Q2533/107 , C12Q2521/501 , C12Q2537/125 , C12Q2525/161 , C12Q2565/519
摘要： An improved method allowing for rapid sensitive and standardized detection of a target nucleic acid from a pathogenic microorganism or virus or normal or abnormal gene in a sample is provided. The method involves hybridizing a target nucleic acid to several non-overlapping oligonucleotide probes that hybridize to adjacent regions in the target nucleic acid, the probes being referred to capture/amplification probes and amplification probes, respectively, in the presence of paramagnetic beads coated with a ligand binding moiety. Through the binding of a ligand attached to one end of the capture/amplification probe and the specific hybridization of portions of the probes to adjacent sequences in the target nucleic acid, a complex comprising the target nucleic acid, the probes and the paramagnetic beads is formed. The probes may then ligated together to form a contiguous ligated amplification sequence bound to the beads, which complex may be denatured to remove the target nucleic acid and unligated probes. Alternatively, separate capture and amplification probes may be used which form continuous full-length or circular probes, and may be directly detected or amplified using a suitable amplification technique, e.g., PCR, RAM or HSAM for detection. The detection of the ligated amplification sequence, either directly or following amplification of the ligated amplification sequence, indicates the presence of the target nucleic acid in a sample. Methods for the detection of the ligated amplification sequence, including hybridization signal amplification method and ramification-extension amplification method, are also provided.
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公开(公告)号：US5942391A
公开(公告)日：1999-08-24
申请号：US690494
申请日：1996-07-31
CPC分类号： C12Q1/686 , C12Q1/6816 , C12Q1/6834 , C12Q1/6862 , C12Q1/689 , Y10T436/143333
摘要： An improved method allowing for rapid sensitive and standardized detection of a target nucleic acid from a pathogenic microorganism or virus or normal or abnormal gene in a sample is provided. The method involves hybridizing a target nucleic acid to several non-overlapping oligonucleotide probes that hybridize to adjacent regions in the target nucleic acid, the probes being referred to capture/amplification probes and amplification probes, respectively, in the presence of paramagnetic beads coated with a ligand binding moiety. Through the binding of a ligand attached to one end of the capture/amplification probe and the specific hybridization of portions of the probes to adjacent sequences in the target nucleic acid, a complex comprising the target nucleic acid, the probes and the paramagnetic beads is formed. The probes may then ligated together to form a contiguous ligated amplification sequence bound to the beads, which complex may be denatured to remove the target nucleic acid and unligated probes. Alternatively, separate capture and amplification probes may be used which form continuous full-length or circular probes, and may be directly detected or amplified using a suitable amplification technique, e.g., PCR, RAM or HSAM for detection. The detection of the ligated amplification sequence, either directly or following amplification of the ligated amplification sequence, indicates the presence of the target nucleic acid in a sample. Methods for the detection of the ligated amplification sequence, including hybridization signal amplification method and ramification-extension amplification method, are also provided.
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3.再颁专利Nucleic acid amplification method hybridization signal amplification method (HSAM) 有权核酸扩增方法杂交信号扩增法（HSAM）公开(公告)号：USRE38442E1
公开(公告)日：2004-02-24
申请号：US09798641
申请日：2001-03-02
申请人： David Y. Zhang , Margaret Brandwein
发明人： David Y. Zhang , Margaret Brandwein
IPC分类号： C12Q170
CPC分类号： C12Q1/6862 , C12Q1/6813 , C12Q1/682 , C12Q1/6834 , C12Q1/6844 , C12Q1/6853 , C12Q1/686 , C12Q1/6865 , G01N33/536 , C12Q2563/131 , C12Q2563/143 , C12Q2565/519
摘要： An improved method allowing for rapid sensitive and standardized detection of a target nucleic acid from a pathogenic microorganism or virus or normal or abnormal gene in a sample is provided. The method involves hybridizing a target nucleic acid to several non-overlapping oligonucleotide probes that hybridize to adjacent regions in the target nucleic acid, the probes being referred to capture/amplification probes and amplifications probes, respectively, in the presence of paramagnetic beads coated with a ligand binding moiety. Through the binding of a ligand attached to one end of the capture/amplification probe and the specific hybridization of portions of the probes to adjacent sequences in the target nucleic acid, a complex comprising the target nucleic acid, the probes and the paramagnetic beads is formed. The probes may then ligated together to form a contiguous ligated amplification sequence bound to the beads, which complex may be denatured to remove the target nucleic acid and unligated probes. Alternatively, separate capture and amplification probes may be used which form continuous full-length or circular probes, and may be directly detected or amplified using a suitable amplification technique, e.g., PCR, RAM or HSAM for detection. The detection of the ligated amplification sequence, either directly or following amplification of the ligated amplification sequence, indicates the presence of the target nucleic acid in a sample. Methods for the detection of the ligated amplification sequence, including hybridization signal amplification method and ramification- extension amplification method, are also provided.
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4.发明授权Nucleic acid amplification method: ramification-extension amplification method (RAM) 有权核酸扩增方法：分枝扩增法（RAM）公开(公告)号：US06855523B2
公开(公告)日：2005-02-15
申请号：US10309438
申请日：2002-12-04
CPC分类号： C12Q1/686 , C12Q1/6816 , C12Q1/6834 , C12Q1/6862 , C12Q1/689 , Y10T436/143333 , C12Q2565/543 , C12Q2563/143 , C12Q2525/185 , C12Q2525/307 , C12Q2523/101 , C12Q2563/131 , C12Q2533/107 , C12Q2521/501 , C12Q2537/125 , C12Q2525/161 , C12Q2565/519
摘要： An improved method allowing for rapid sensitive and standardized detection of a target nucleic acid from a pathogenic microorganism or virus or normal or abnormal gene in a sample is provided. The method involves hybridizing a target nucleic acid to several non-overlapping oligonucleotide probes that hybridize to adjacent regions in the target nucleic acid, the probes being referred to capture/amplification probes and amplification probes, respectively, in the presence of paramagnetic beads coated with a ligand binding moiety. Through the binding of a ligand attached to one end of the capture/amplification probe and the specific hybridization of portions of the probes to adjacent sequences in the target nucleic acid, a complex comprising the target nucleic acid, the probes and the paramagnetic beads is formed. The probes may then ligated together to form a contiguous ligated amplification sequence bound to the beads, which complex may be denatured to remove the target nucleic acid and unligated probes. Alternatively, separate capture and amplification probes may be used which form continuous full-length or circular probes, and may be directly detected or amplified using a suitable amplification technique, e.g., PCR, RAM or HSAM for detection. The detection of the ligated amplification sequence, either directly or following amplification of the ligated amplification sequence, indicates the presence of the target nucleic acid in a sample. Methods for the detection of the ligated amplification sequence, including hybridization signal amplification method and ramification-extension amplification method, are also provided.
摘要翻译： 提供了一种允许从病原微生物或病毒或样品中的正常或异常基因对靶核酸进行快速敏感和标准化检测的改进方法。 该方法包括将目标核酸杂交到与靶核酸中的相邻区域杂交的几个非重叠寡核苷酸探针杂交，探针分别参照捕获/扩增探针和扩增探针，在涂覆有 配体结合部分。 通过连接到捕获/扩增探针的一端的配体的结合和探针的部分与目标核酸中的相邻序列的特异性杂交，形成包含靶核酸，探针和顺磁珠的复合物 。 然后可以将探针连接在一起以形成与珠结合的连续连接的扩增序列，该复合物可能被变性以除去靶核酸和未结合的探针。 或者，可以使用分离的捕获和扩增探针，其形成连续的全长或圆形探针，并且可以使用合适的扩增技术（例如PCR，RAM或HSAM）直接检测或扩增用于检测。 连接的扩增序列的检测直接或连续扩增序列的扩增后，表明靶核酸在样品中的存在。 还提供了检测连接扩增序列的方法，包括杂交信号扩增方法和分枝扩增扩增方法。
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5.发明授权Nucleic acid amplification method hybridization signal amplification method (HSAM) 失效核酸扩增方法杂交信号扩增法（HSAM）
公开(公告)号：US5876924A
公开(公告)日：1999-03-02
申请号：US690495
申请日：1996-07-31
申请人： David Y. Zhang , Margaret Brandwein
发明人： David Y. Zhang , Margaret Brandwein
CPC分类号： C12Q1/6844 , C12Q1/6813 , C12Q1/682 , C12Q1/6834 , C12Q1/686 , C12Q1/6862 , C12Q1/6865
摘要： An improved method allowing for rapid sensitive and standardized detection of a target nucleic acid from a pathogenic microorganism or virus or normal or abnormal gene in a sample is provided. The method involves hybridizing a target nucleic acid to several non-overlapping oligonucleotide probes that hybridize to adjacent regions in the target nucleic acid, the probes being referred to capture/amplification probes and amplification probes, respectively, in the presence of paramagnetic beads coated with a ligand binding moiety. Through the binding of a ligand attached to one end of the capture/amplification probe and the specific hybridization of portions of the probes to adjacent sequences in the target nucleic acid, a complex comprising the target nucleic acid, the probes and the paramagnetic beads is formed. The probes may then ligated together to form a contiguous ligated amplification sequence bound to the beads, which complex may be denatured to remove the target nucleic acid and unligated probes. Alternatively, separate capture and amplification probes may be used which form continuous full-length or circular probes, and may be directly detected or amplified using a suitable amplification technique, e.g., PCR, RAM or HSAM for detection. The detection of the ligated amplification sequence, either directly or following amplification of the ligated amplification sequence, indicates the presence of the target nucleic acid in a sample. Methods for the detection of the ligated amplification sequence, including hybridization signal amplification method and ramification- extension amplification method, are also provided.
摘要翻译： 提供了一种允许从病原微生物或病毒或样品中的正常或异常基因对靶核酸进行快速敏感和标准化检测的改进方法。 该方法包括将目标核酸杂交到与靶核酸中的相邻区域杂交的几个非重叠寡核苷酸探针杂交，探针分别参照捕获/扩增探针和扩增探针，在涂覆有 配体结合部分。 通过连接到捕获/扩增探针的一端的配体的结合和探针的部分与目标核酸中的相邻序列的特异性杂交，形成包含靶核酸，探针和顺磁珠的复合物 。 然后可以将探针连接在一起以形成与珠结合的连续连接的扩增序列，该复合物可能被变性以除去靶核酸和未结合的探针。 或者，可以使用分离的捕获和扩增探针，其形成连续的全长或圆形探针，并且可以使用合适的扩增技术（例如PCR，RAM或HSAM）直接检测或扩增用于检测。 连接的扩增序列的检测直接或连续扩增序列的扩增后，表明靶核酸在样品中的存在。 还提供了检测连接扩增序列的方法，包括杂交信号扩增方法和分枝扩增扩增方法。
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