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    • 4. 发明授权
    • Sequence-specific methods for homogeneous, real-time detection of lamp products
    • 灯具产品均匀,实时检测的序列特异性方法
    • US09315863B2
    • 2016-04-19
    • US13505598
    • 2010-11-04
    • James G. Nadeau
    • James G. Nadeau
    • C12Q1/68
    • C12Q1/6865C12Q2525/307
    • Presented herein are methods and compositions for generating sequence-specific, secondary amplification products during Loop-mediated Isothermal Amplification (LAMP). Conventional LAMP produces a preponderance of high molecular weight DNA structures concatenated into self-complementary hairpins, which are not amenable to detection by routine probe-based hybridization methods, making multiplex detection of two or more targets or sequence variants in closed-tube formats extremely difficult. Provided herein, for example, are methods for generating secondary LAMP products bearing a fragment of the original target sequence embedded within low-molecular weight products that are devoid of competitive hairpin structures, the lack of which enhances probe-based detection of target sequences. These secondary products can, for example, be produced in real-time, during the LAMP process, and can provide the option of detecting multiple target sequences within a single tube using, e.g., a homogenous, real-time fluorescence format.
    • 本文提出了在环介导的等温扩增(LAMP)期间产生序列特异性二级扩增产物的方法和组合物。 常规的LAMP产生连接到自身互补发夹中的高分子量DNA结构的优势,其不适于通过常规基于探针的杂交方法的检测,使得封闭管形式中的两个或更多个目标或序列变体的多重检测非常困难 。 例如,本文提供的是产生具有嵌入低分子量产物中的原始靶序列的片段的次级LAMP产物的方法,所述片段不含有竞争力的发夹结构,其缺乏可增强靶序列的基于探针的检测。 这些次要产物可以例如在LAMP过程期间实时产生,并且可以提供使用例如均匀的实时荧光格式在单个管内检测多个靶序列的选项。
    • 7. 发明授权
    • Simultaneous amplification of multiple targets
    • 同时放大多个目标
    • US5422252A
    • 1995-06-06
    • US73197
    • 1993-06-04
    • George T. WalkerJames G. NadeauMichael C. Little
    • George T. WalkerJames G. NadeauMichael C. Little
    • G01N33/566C07H21/04C12N15/09C12P19/34C12Q1/68G11B5/00G11B5/09G11B5/187G11B5/31G11B5/455G11B5/465G11B19/04C12Q1/70
    • C12Q1/6853
    • Methods for multiplex amplification of target nucleic acid sequences using a single pair of primers. Defined sequences are appended to the ends of multiple target sequences as part of the amplification reaction so that no steps in addition to amplification are required. The target sequences with the appended defined sequences need not be isolated prior to amplification. In one embodiment for coamplification of two target sequences, a sequence corresponding to a terminal segment of the first target sequence is appended to one end of the second target sequence and a sequence corresponding to a terminal segment of the second target sequence is appended to one end of the first target sequence. Amplification of the two targets then requires only a single pair of primers. Alternatively, a single defined sequence may be appended to the 5' and 3' ends of any number of selected targets. All such modified target sequences may then be amplified using a single pair of primers which hybridize to the defined end-sequences.
    • 使用单对引物多重扩增靶核酸序列的方法。 定义的序列作为扩增反应的一部分附加到多个靶序列的末端,因此不需要扩增步骤。 具有附加定义序列的靶序列不需要在扩增前分离。 在两个目标序列的共放大的一个实施例中,对应于第一目标序列的末端片段的序列被附加到第二目标序列的一端,并且与第二目标序列的末端片段相对应的序列被附加到一端 的第一个靶序列。 两个靶标的扩增只需要一对引物即可。 或者,单个定义的序列可以附加到任意数目的所选目标的5'和3'端。 然后可以使用与定义的末端序列杂交的单对引物扩增所有这些修饰的靶序列。
    • 10. 发明授权
    • Simultaneous amplification of multiple targets
    • 同时放大多个目标
    • US5624825A
    • 1997-04-29
    • US451313
    • 1995-05-26
    • George T. WalkerJames G. NadeauMichael C. Little
    • George T. WalkerJames G. NadeauMichael C. Little
    • G01N33/566C07H21/04C12N15/09C12P19/34C12Q1/68G11B5/00G11B5/09G11B5/187G11B5/31G11B5/455G11B5/465G11B19/04
    • C12Q1/6853
    • Methods for multiplex amplification of target nucleic acid sequences using a single pair of primers. Defined sequences are appended to the ends of multiple target sequences as part of the amplification reaction so that no steps in addition to amplification are required. The target sequences with the appended defined sequences need not be isolated prior to amplification. In one embodiment for coamplification of two target sequences, a sequence corresponding to a terminal segment of the first target sequence is appended to one end of the second target sequence and a sequence corresponding to a terminal segment of the second target sequence is appended to one end of the first target sequence. Amplification of the two targets then requires only a single pair of primers. Alternatively, a single defined sequence may be appended to the 5' and 3' ends of any number of selected targets. All such modified target sequences may then be amplified using a single pair of primers which hybridize to the defined end-sequences.
    • 使用单对引物多重扩增靶核酸序列的方法。 定义的序列作为扩增反应的一部分附加到多个靶序列的末端,因此不需要扩增步骤。 具有附加定义序列的靶序列不需要在扩增前分离。 在两个目标序列的共放大的一个实施例中,对应于第一目标序列的末端片段的序列被附加到第二目标序列的一端,并且与第二目标序列的末端片段相对应的序列被附加到一端 的第一个靶序列。 两个靶标的扩增只需要一对引物即可。 或者,单个定义的序列可以附加到任意数目的所选目标的5'和3'端。 然后可以使用与定义的末端序列杂交的单对引物扩增所有这些修饰的靶序列。