公开(公告)号：US06355709B1

公开(公告)日：2002-03-12

申请号：US08950926

申请日：1997-10-15

申请人： Ramakrishna S. Madabhushi ,  Steven M. Menchen ,  J. William Efcavitch ,  Paul D. Grossman

发明人： Ramakrishna S. Madabhushi ,  Steven M. Menchen ,  J. William Efcavitch ,  Paul D. Grossman

IPC分类号： C08K516

CPC分类号： G01N27/44752 ,  G01N27/44747

摘要： The invention provides uncharged water-soluble silica-adsorbing polymers for suppressing electroendoosmotic flow and to reduce analyte-wall interactions in capillary electrophoresis. In one aspect of the invention, one or more of such polymers are employed as components of a separation medium for the separation of biomolecules, such as polynucleotides, polysaccharides, proteins, and the like, by capillary electrophoresis. Generally, such polymers are characterized by (i) water solubility over the temperature range between about 20° C. to about 50° C., (ii) concentration in a separation medium in the range between about 0.001% to about 10% (weight/volume), (iii) molecular weight in the range of about 5×103 to about 1×106 daltons, and (iv) absence of charged groups in an aqueous medium having pH in the range of about 6 to about 9. In one embodiment, polymers of the invention are selected from the group consisting of polylactams, such as polyvinylpyrrolidone; N,N-disubstituted polyacrylamides; and N-substituted polyacrylamides. In accordance with the method of the invention, a sufficient amount of polymer adsorbs to the capillary surface to establish a zone of high viscosity that shields the analyte from the wall and impedes the movement of an electrical double layer under an electric field.

公开(公告)号：US5989871A

公开(公告)日：1999-11-23

申请号：US800641

申请日：1997-02-14

申请人： Paul D. Grossman ,  Steven Fung ,  Steven M. Menchen ,  Sam Lee Woo ,  Emily S. Winn-Deen

发明人： Paul D. Grossman ,  Steven Fung ,  Steven M. Menchen ,  Sam Lee Woo ,  Emily S. Winn-Deen

IPC分类号： C12N15/09 ,  B01D57/02 ,  C12Q1/68 ,  G01N27/447 ,  G01N30/88 ,  G01N33/566 ,  C12P19/34 ,  C07H21/04

CPC分类号： C12Q1/6869 ,  C12Q1/6816 ,  C12Q1/6827 ,  C12Q1/6858 ,  C12Q1/686 ,  C12Q1/6862 ,  C12Q1/6818 ,  Y10T436/143333

摘要： Method and composition for detecting one or more selected polynucleotide regions in a target polynucleotide. In the method, a mixture of sequence-specific probes are reacted with the target polynucleotide under hybridization conditions, and the hybridized probes are treated to selectively modify those probes which are bound to the target polynucleotide in a base-specific manner. The resulting labeled probes include a polymer chain which imparts to each different-sequence probe, a distinctive ratio of charge/translational frictional drag, and a detectable label. The labeled probes are fractionated by electrophoresis in a non-sieving matrix, and the presence of one or more selected sequences in the target polynucleotide are detected according to the observed electrophoretic migration rates of the labeled probes in a non-sieiving medium.

摘要翻译： 用于检测靶多核苷酸中的一个或多个选择的多核苷酸区域的方法和组合物。 在该方法中，将序列特异性探针的混合物与靶多核苷酸在杂交条件下反应，并对杂交的探针进行处理以选择性地修饰以碱基特异性方式结合靶多核苷酸的那些探针。 所得到的标记探针包括赋予每个不同序列探针的聚合物链，电荷/平移摩擦阻力的独特比例和可检测标记。 标记的探针通过在非筛选基质中的电泳进行分级，并且根据在非筛选培养基中标记的探针的观察到的电泳迁移速率来检测靶多核苷酸中一个或多个选定序列的存在。

公开(公告)号：US5374527A

公开(公告)日：1994-12-20

申请号：US3968

申请日：1993-01-21

申请人： Paul D. Grossman

发明人： Paul D. Grossman

IPC分类号： G01N33/483 ,  C12M1/00 ,  C12N15/09 ,  C12Q1/68 ,  G01N27/447 ,  G01N33/50 ,  C12P19/34 ,  G01N33/00

CPC分类号： C12Q1/6869 ,  Y10T436/143333

摘要： A DNA sequencing method for use in sequencing a DNA target sequence up to 300 bases, preferably up to 500 bases or greater in length, by electrophoretically separating a mixture of single-stranded DNA sequencing fragments in a capillary tube. The method employs an aqueous denaturing solution comprising between about 4 and about 7 weight percent linear polyacrylamide molecules having an average molecular weight of between about 20 and about 100 kDa. The low-viscosity of the solution allows rapid loading and reloading of such solution into the capillary tube.

摘要翻译： 一种DNA测序方法，用于通过在毛细管中电泳分离单链DNA测序片段的混合物，将DNA靶序列测序长达300个碱基，优选长达500个碱基或更大。 该方法使用包含约4至约7重量％的平均分子量为约20至约100kDa的线性聚丙烯酰胺分子的水性变性溶液。 溶液的低粘度允许将这种溶液快速加载和重新加载到毛细管中。

公开(公告)号：US5346999A

公开(公告)日：1994-09-13

申请号：US328471

申请日：1989-03-24

申请人： Guy R. Cathcart ,  Paul D. Grossman ,  P. Eric Mayrand ,  Eric S. Nordman ,  Norman M. Whiteley

发明人： Guy R. Cathcart ,  Paul D. Grossman ,  P. Eric Mayrand ,  Eric S. Nordman ,  Norman M. Whiteley

IPC分类号： B01J19/00 ,  C12M1/00 ,  C12N9/06 ,  C12N15/10 ,  C40B60/14 ,  C07H23/00

CPC分类号： C12N9/0012 ,  B01J19/0046 ,  C12M47/06 ,  C12M47/12 ,  C12N15/1003 ,  B01J2219/00286 ,  B01J2219/00308 ,  B01J2219/00331 ,  B01J2219/00389 ,  B01J2219/00488 ,  B01J2219/00689 ,  B01J2219/00695 ,  C40B60/14 ,  Y10T436/25125 ,  Y10T436/25375

摘要： An automated apparatus is provided which implements a new method of extracting and purifying nucleic acids from cells without the use of centrifugation. In the method, a lysate is created by treating the cells with proteinase K in the presence of a lysis buffer having a high concentration of a salt. The lysate is mixed with a phenol-based solvent system, thereby creating an emulsion. The emulsion is heated to promote phase separation. Similarly, the rate of phase separation is also enhanced by increasing the surface area of the emulsion. Once the phase separation is complete, the lower organic phase is removed and the upper aqueous phase is repeatedly extracted with the phenol-based solvent a preselected number of times, and is finally extracted using chloroform. The remaining aqueous phase is then dialyzed to further purify and concentrate the nucleic acid solution. Two preferred embodiments of apparatus are presented to accomplish this extraction.

摘要翻译： 提供了一种自动化装置，其实现了从细胞中提取和纯化核酸而不使用离心的新方法。 在该方法中，通过在具有高浓度盐的裂解缓冲液存在下用蛋白酶K处理细胞产生裂解物。 将裂解物与苯酚基溶剂体系混合，从而产生乳液。 将乳液加热以促进相分离。 类似地，通过增加乳液的表面积也增加了相分离速率。 一旦相分离完成，除去较低的有机相，并用苯酚类溶剂预选次数反复提取上部水相，最后用氯仿萃取。 然后将剩余的水相透析以进一步净化并浓缩核酸溶液。 呈现装置的两个优选实施例以完成该提取。