Compositions, methods, and kits for isolating nucleic acids...

Details Compositions, methods, and kits for isolating nucleic acids... Compositions, methods, and kits for isolating nucleic acids...

2001-11-28

2004-07-13

Riley, Jezia (Department: 1637)

Chemistry: molecular biology and microbiology

Measuring or testing process involving enzymes or...

Involving nucleic acid

C435S243000, C536S023100, C536S026420, C536S027120

Reexamination Certificate

active

06762027

ABSTRACT:

FIELD OF THE INVENTION
The invention relates to compositions and methods for releasing and isolating nucleic acids from biological samples, including whole tissue. The invention also provides kits for isolating and/or releasing nucleic acids from biological samples.
BACKGROUND OF THE INVENTION
Current methods for isolating nucleic acids from biological samples may include macerating tissues, lysing cells, and inactivating nucleases using chaotropic salts, such as guanidine hydrochloride or guanidinium thiocyanate, and a nonionic detergent. The released nucleic acids then may be selectively precipitated from solution.
Various chemical disruption methods, using detergents, chaotropes, proteases, bile salts, organic solvents, and harsh acidic or basic conditions have been employed to macerate tissue and release nucleic acid. The nucleic acid obtained using such methods may be degraded due to long incubation times or exposure to harsh conditions.
SUMMARY OF THE INVENTION
In certain embodiments, compositions are provided for releasing nucleic acids from a biological sample. In certain embodiments, the compositions include at least one cationic surfactant, at least one protease, and a buffer.
In certain embodiments, methods for releasing nucleic acids from a biological sample are provided. In certain embodiments, methods for isolating nucleic acids from a biological sample are provided. In certain embodiments, these methods include combining the sample with at least one cationic surfactant, at least one protease, and a buffer to form a reaction composition. In certain embodiments, the reaction composition further comprises a second surfactant. In certain embodiments, the reaction composition further comprises a salt. In certain embodiments, the composition is incubated under appropriate conditions to release the nucleic acid from the biological sample. In certain embodiments, the released nucleic acid is then isolated.
In certain embodiments, kits for isolating and/or releasing nucleic acid from biological samples are also provided. In certain embodiments, the invention provides kits comprising at least one cationic surfactant and at least one protease. In certain embodiments, kits further comprise a second surfactant, a salt, organic extraction agent(s), organic precipitating agent(s), solubilizing agents(s), nucleic acid-binding solid phase(s), or combinations of these components.
According to certain embodiments, a method of obtaining nucleic acid from a biological sample and binding the nucleic acid to a solid phase is provided. In certain embodiments, this method comprises contacting the biological sample with a disrupting buffer, wherein the disrupting buffer comprises a protease and a cationic surfactant; and binding the nucleic acid to a solid phase. In certain embodiments, the method comprises contacting the biological sample with a disrupting buffer, wherein the disrupting buffer comprises a protease and a cationic surfactant; substantially neutralizing the cationic surfactant; and binding the nucleic acid to a solid phase.
According to certain embodiments, a kit is provided, comprising a protease, a cationic surfactant, and a second surfactant, wherein the second surfactant permits the binding of nucleic acid to a solid phase in the presence of the protease and cationic surfactant.
According to certain embodiments, a kit for obtaining nucleic acid from a biological sample is provided, comprising a protease; a cationic surfactant; a non-ionic surfactant, wherein the non-ionic surfactant permits the binding of nucleic acid to a solid phase in the presence of the protease and cationic surfactant; and a high salt buffer.


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