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1. 发明授权

US5707808A Optical selection and collection of DNA fragments 失效
公开(公告)号：US5707808A
公开(公告)日：1998-01-13
申请号：US632743
申请日：1996-04-15
申请人： Mary C. Roslaniec , John C. Martin , James H. Jett , L. Scott Cram
发明人： Mary C. Roslaniec , John C. Martin , James H. Jett , L. Scott Cram
IPC分类号： C12N15/10 , C12N15/12 , C12Q1/68 , G01N15/14 , G01N21/64
CPC分类号： G01N15/1459 , C12N15/1006 , C12Q1/68 , G01N2015/0038 , G01N2015/149 , Y10T436/143333
摘要： Optical selection and collection of DNA fragments. The present invention includes the optical selection and collection of large (>.mu.g) quantities of clonable, chromosome-specific DNA from a sample of chromosomes. Chromosome selection is based on selective, irreversible photoinactivation of unwanted chromosomal DNA. Although more general procedures may be envisioned, the invention is demonstrated by processing chromosomes in a conventional flow cytometry apparatus, but where no droplets are generated. All chromosomes in the sample are first stained with at least one fluorescent analytic dye and bonded to a photochemically active species which can render chromosomal DNA unclonable if activated. After passing through analyzing light beam(s), unwanted chromosomes are irradiated using light which is absorbed by the photochemically active species, thereby causing photoinactivation. As desired chromosomes pass this photoinactivation point, the inactivating light source is deflected by an optical modulator; hence, desired chromosomes are not photoinactivated and remain clonable. The selection and photoinactivation processes take place on a microsecond timescale. By eliminating droplet formation, chromosome selection rates 50 times greater than those possible with conventional chromosome sorters may be obtained. Thus, usable quantities of clonable DNA from any source thereof may be collected.

2. 发明授权

US4784737A Electromicroinjection of particles into living cells 失效
标题翻译：将微粒体注入活细胞
公开(公告)号：US4784737A
公开(公告)日：1988-11-15
申请号：US853823
申请日：1986-04-18
申请人： F. Andrew Ray , L. Scott Cram , William R. Galey
发明人： F. Andrew Ray , L. Scott Cram , William R. Galey
IPC分类号： A61N1/30 , C12M3/00 , C12N13/00 , C12N15/89 , B01D57/02 , A61B17/36 , C12N15/00
CPC分类号： C12N15/89 , A61N1/306 , C12M35/02 , C12N13/00
摘要： Method and apparatus for introducing particles into living cells. Fluorescently-stained human chromosomes are introduced into cultured, mitotic Chinese hamster cells using electromicroinjection. The recipient cells frequently survived the physiological perturbation imposed by a successful chromosome injection. Successfully injected recipient cells maintained viability as evidenced by their ability to be expanded.The technique relies on the surface charge of fluorescently stained chromosomes and their ability to be attracted and repelled to and from the tip of a micropipette. The apparatus includes a micropipette having a tip suitable for piercing the membrane of a target cell and an electrode inserted into the lumen thereof. The target cells and suspended particles are located in an electrically conducted solution, and the lumen of the micropipette is filled with an electrically conducting solution which contacts the electrode located therein. A second electrode is also located in the conducting solution containing the target cells and particles. Voltages applied to the electrode within the micropipette attract the particles to the region of the tip thereof. The particles adhere to the surface of the micropipette with sufficient force that insertion of the micropipette tip and attached particle through the membrane of a target cell will not dislodge the particle. By applying a voltage having the opposite polarity of the attraction voltage, the particles are expelled from the micropipette to which is then withdrawn from the cell body.
摘要翻译：将颗粒引入活细胞的方法和装置。荧光染色的人类染色体使用电喷雾引入培养的有丝分裂的中国仓鼠细胞。受体细胞经常由成功的染色体注射施加的生理扰动中存活。成功注射受体细胞保持活力，这是由于其扩展能力所证明。该技术依赖于荧光染色体的表面电荷及其吸附和排斥到微量移液管尖端的能力。该装置包括具有尖端的微量移液管，该尖端适于刺穿靶细胞的膜和插入其内腔中的电极。靶细胞和悬浮颗粒位于导电溶液中，并且微量移液管的内腔填充有与位于其中的电极接触的导电溶液。第二电极也位于含有靶细胞和颗粒的导电溶液中。施加到微量移液管内的电极的电压将颗粒吸引到其尖端的区域。颗粒以足够的力粘附到微量吸管的表面，使得微量吸头和附着的颗粒通过靶细胞的膜插入不会移除颗粒。通过施加具有与吸引电压相反的极性的电压，颗粒从微量移液管排出，然后将其从细胞体中取出。

3. 发明授权

US5879625A Optical selection and collection of DNA fragments 失效
标题翻译： DNA片段的光学选择和收集
公开(公告)号：US5879625A
公开(公告)日：1999-03-09
申请号：US951955
申请日：1997-10-17
申请人： Mary C. Roslaniec , John C. Martin , James H. Jett , L. Scott Cram
发明人： Mary C. Roslaniec , John C. Martin , James H. Jett , L. Scott Cram
IPC分类号： C12N15/10 , C12N15/12 , C12Q1/68 , G01N15/14 , G01N1/14
CPC分类号： G01N15/1459 , C12N15/1006 , C12Q1/68 , G01N2015/0038 , G01N2015/149 , Y10T436/143333
摘要： Optical selection and collection of DNA fragments. The present invention includes the optical selection and collection of large (>.mu.g) quantities of clonable, chromosome-specific DNA from a sample of chromosomes. Chromosome selection is based on selective, irreversible photoinactivation of unwanted chromosomal DNA. Although more general procedures may be envisioned, the invention is demonstrated by processing chromosomes in a conventional flow cytometry apparatus, but where no droplets are generated. All chromosomes in the sample are first stained with at least one fluorescent analytic dye and bonded to a photochemically active species which can render chromosomal DNA unclonable if activated. After passing through analyzing light beam(s), unwanted chromosomes are irradiated using light which is absorbed by the photochemically active species, thereby causing photoinactivation. As desired chromosomes pass this photoinactivation point, the inactivating light source is deflected by an optical modulator; hence, desired chromosomes are not photoinactivated and remain clonable. The selection and photoinactivation processes take place on a microsecond timescale. By eliminating droplet formation, chromosome selection rates 50 times greater than those possible with conventional chromosome sorters may be obtained. Thus, usable quantities of clonable DNA from any source thereof may be collected.
摘要翻译： DNA片段的光学选择和收集。本发明包括从染色体样品中大量（>μg）量的可克隆，染色体特异性DNA的光学选择和收集。染色体选择基于不需要的染色体DNA的选择性不可逆光激活。尽管可以设想更一般的程序，但是通过在常规流式细胞术装置中处理染色体但是不产生液滴来证明本发明。样品中的所有染色体首先用至少一种荧光分析染料染色，并与光化学活性物质结合，如果激活可使染色体DNA不可克隆。在通过分析光束之后，使用被光化学活性物质吸收的光照射不想要的染色体，从而引起光激活。根据需要，染色体通过该光激活点，灭活光源被光学调制器偏转; 因此，期望的染色体不是光激活的并且保持克隆。选择和光激活过程在微秒的时间尺度上进行。通过消除液滴形成，可以获得比常规染色体分选机可能的染色体选择率高50倍的染色体选择率。因此，可以收集来自其任何来源的可克隆DNA的可用量。

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