Transgenomic, Inc.

David W Togami

Douglas T Gjerde

Robert M Haefele

Improved liquid chromatography systems having components made of titanium, stainless steel, or organic polymeric material are useful in the separation of polynucleotide fragments, particularly large fragments of double-stranded polynucleotides, by Matched Ion Polynucleotide Chromatography (MIPC). The titanium, stainless steel, or polymeric components are treated so that they do not release multivalent cations into aqueous solutions flowing through the chromatography system. Alternatively, or in addition to utilizing materials made of the components listed above, a multivalent cation capture resin placed upstream of the separation column can be employed to remove multivalent ions from the system. The multivalent cation capture resin can be contained in a guard disk, a guard column, or a guard cartridge. Novel methods for separating mixtures of polynucleotide fragments into fractions based on their molecular weight by Matched Ion Polynucleotide Chromatography and slalom chromatography utilize the liquid chromatographic systems described above.

System and method for performing polynucleotide separations using liquid chromatography

The invention recognizes the deleterious effects of trace, and even undetectable amounts of multivalent cations on the separation of mixtures of polynucleotides, especially double stranded polynucleotides, and provides an improved method for separating such mixtures on wide pore, non-polar separation media by eliminating multivalent cations from the all aspects of the separation process. This is accomplished by using components in the separation process which are materials which do not release metal cations. In addition, the use of cation capture resins and other methods to remove residual traces of multivalent cations from eluting solvents, sample solutions, separation media, and system components is described. It is also important to remove any traces or organic contaminants from solvents solutions and system parts. Taking similar steps to remove residual traces of multivalent cations and organic impurities from the separation process, the invention may also be used in a batch process to separate mixtures of polynucleotide fragments.

Process for performing polynucleotide separations

System and column for performing polynucleotide separations using liquid chromatography

Method for performing polynucleotide separations using liquid chromatography

Apparatus for performing polynucleotide separations using liquid

Method for performing polynucleotide separations using liquid

Improved liquid chromatography systems having components made of titanium, coated stainless steel, or organic polymeric material are useful in the separation of nucleic acid fragments, particularly large fragments of double-stranded nucleic acids, by ion pairing reverse phase chromatography. The titanium, coated stainless steel, or polymeric components do not release multivalent cations into aqueous solutions flowing through the chromatography system. Alternatively, or in addition to utilizing materials made of the components listed above, a multivalent cation capture resin placed upstream of the separation column may be employed to remove multivalent ions from the system. The multivalent cation capture resin may be contained in a guard disk, a guard column, or a guard cartridge. Novel methods for separating mixtures of nucleic acid fragments into fractions based on their molecular weight by ion pairing reverse phase chromatography and slalom chromatography utilize the liquid chromatographic systems described above.

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San Jose, CA, United States of America

David W Togami