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    Methods for amplification of nucleic acids utilizing clamp oligonucleotides
    2.
    发明授权
    Methods for amplification of nucleic acids utilizing clamp oligonucleotides 有权

    公开(公告)号：US10202629B2

    公开(公告)日：2019-02-12

    申请号：US14773365

    申请日：2014-03-14

    申请人： Lyle J. Arnold ,  Norman C. Nelson

    发明人： Lyle J. Arnold

    IPC分类号： C12P19/34 ,  C12Q1/68 ,  C12Q1/6844

    摘要： The present invention provides methods of amplifying a target nucleic acid utilizing a clamp oligonucleotide comprising a first target-binding region on the 3′-terminus and a second target-binding region on the 5′-terminus and tether region in between. The tether region may comprise a variety of user-defined sequences or elements that allow for further manipulation of the target nucleic acid. Such as, for example, capture followed by amplification, identification and/or sequencing. The target-binding regions bind to the target nucleic acid, the 3′-terminus functions as a primer to initiate extension across the target nucleic acid sequence and ligation of the gap results in formation of a circularized nucleic acid. This circular template can be used in a variety of processes, including amplification and sequencing.

    Methods for amplification of nucleic acids utilizing hairpin loop or duplex primers
    3.
    发明授权
    Methods for amplification of nucleic acids utilizing hairpin loop or duplex primers 有权

    公开(公告)号：US10066262B2

    公开(公告)日：2018-09-04

    申请号：US14999978

    申请日：2016-07-21

    申请人： Lyle J. Arnold

    发明人： Lyle J. Arnold

    IPC分类号： C12P19/34 ,  C12Q1/6853 ,  C12N15/10

    摘要： The present invention provides methods of amplifying a target nucleic acid utilizing duplex primer. The first strand of the primer comprises a random nucleotide sequence of about 6 to about 9 nucleotides in length that is able to hybridize to the target nucleic acid and a tag sequence. The second strand of the primer comprises a sequence complementary to the tag sequence allowing the primer to form a duplex and the ability to bind the tag sequence of the product nucleic acid for further amplification. The resulting nucleic acid produced contains tag sequences on both the 3′- and 5′-termini.

    Methods for amplification of nucleic acids on solid support
    5.
    发明授权
    Methods for amplification of nucleic acids on solid support 有权

    公开(公告)号：US10174352B2

    公开(公告)日：2019-01-08

    申请号：US14773362

    申请日：2014-03-14

    申请人： Lyle J. Arnold ,  Norman C. Nelson

    发明人： Lyle J. Arnold ,  Norman C. Nelson

    IPC分类号： C12Q1/68 ,  C12P19/34 ,  C12Q1/6853 ,  C12Q1/6865 ,  C07H21/02

    摘要： The present invention provides methods for amplifying a nucleic acid from a sample containing a mixture of nucleic acids utilizing a solid support. Methods are provided utilizing user-defined primer oligonucleotides for directional amplification that assists in further manipulation of the target nucleic acid, such as sequencing. Methods are also provided utilizing blocker and displacer oligonucleotides for generating amplified target nucleic acids of defined length. One of these methods provides a first oligonucleotide and a second oligonucleotide affixed to a solid support or separate solid supports. The first oligonucleotide is blocked to prevent extension from the 3′-terminus and has a sequence complementary to a first portion of a target nucleic acid. The second oligonucleotide has a sequence that is identical to a second portion of the target nucleic acid. In this method, a sample is applied to the solid support and the target nucleic acid within the sample binds said first oligonucleotide. The solid support is then washed to remove unbound nucleic acids. A primer sequence containing a target binding region and a polymerase promoter sequence is then annealed to the bound target nucleic acid and extended producing a first duplex nucleic acid. The target sequence is then removed leaving a first nucleic acid that can now bind the second oligonucleotide. The second oligonucleotide is extended to produce a second duplex nucleic acid that contains a second nucleic acid. The second nucleic acid is then amplified by adding a polymerase.

    Methods for immobilizing target nucleic acids utilizing combinatorial capture probes
    9.
    发明授权
    Methods for immobilizing target nucleic acids utilizing combinatorial capture probes 有权

    公开(公告)号：US10179931B2

    公开(公告)日：2019-01-15

    申请号：US15652372

    申请日：2017-07-18

    申请人： Lyle J. Arnold, Jr.

    发明人： Lyle J. Arnold, Jr.

    IPC分类号： C12P19/34 ,  C12Q1/6834

    摘要： The present invention provides methods for immobilizing target nucleic acids on a solid support utilizing combinatorial capture probe pairs. These pairs contain first and second capture oligonucleotides that each comprise a target binding region, a capture region and a stem region positioned between the target binding and capture regions. The target binding regions comprise nucleic acid sequences that allow them to hybridize to adjacent regions on the target nucleic acid. The stem regions have nucleic acid sequences that are complementary to each other and the capture regions each comprise a sequence that when positioned adjacent to one another produce a combined nucleic acid sequence that is complementary to a portion of an oligonucleotide bound to a solid support. When the first and second capture oligonucleotides are annealed to the target nucleic acid, the stem regions are brought together allowing them to hybridize, which in turn brings the capture regions together to produce a combined nucleic acid sequence. This combined nucleic acid sequence is then able to hybridize to the oligonucleotide bound to the solid support thereby immobilizing the target nucleic acid.