Abstract This invention relates to novel methods for the release of nucleic acids from cells in complex biological samples or specimens to prepare and make available the nucleic acid material present for a hybridization assay or for extraction. Novel methods for hybridization of nucleic acids are also presented. In particular methods are described for isolating nucleic acid from a sample containing a complex biological mixture of nucleic acid and non-nucleic acids wherein the sample is combined with a hybridization medium comprising a lactam which promotes and enables nucleic acid pairing when complementary nucleic acid is introduced. The lactam is preferably about 5 to about 70% of the hybridization medium and is most preferably 2-pyrrolidone, N-ethyl-2-pyrrolidone, N-cyclohexyl-2-pyrrolidone, N-dodecyl-2-pyrrolidone, N-methyl-2-pyrrolidone, N-hydroxyethyl-2-pyrrolidone, N-methyl-2-piperidone, 2-ε-caprolactam, N-methyl-2-caprolactam, 2-piperidone or N-(4-hydroxybenzyl)pyrrolidone.
A composition for simultaneously lysing cells and hybridizing released endogenous nucleic acid, comprising: a buffer solution; at least one monomeric lactam at a concentration of about 5% to about 70% of the total volume of the buffer solution; and a detergent at a concentration of about 0.05% to about 0.5%.
Claim 2
Dependent Claim
50 words
A composition for simultaneously lysing cells and hybridizing released endogenous nucleic acid, comprising: a buffer solution; at least one monomeric lactam at a concentration of about 5% to about 70% of the total volume of the buffer solution; and a detergent at a concentration of about 0.05% to about 0.5%.
Claim 3
Dependent Claim
10 words
The composition of claim 1 further comprising a chaotropic agent.
Claim 4
Dependent Claim
14 words
The composition of claim 1 further comprising about 1 to about 10 mM EDTA.
Claim 5
Dependent Claim
11 words
The composition of claim 1 further comprising a volume exclusion agent.
Claim 6
Dependent Claim
58 words
A kit for simultaneously lysing cells and hybridizing released endogenous nucleic acid, comprising at least one vial containing a composition comprising: a buffer solution; at least one monomeric lactam at a concentration of about 5% to about 70% of the total volume of the buffer solution; and a detergent at a concentration of about 0.05% to about 0.5%.
Claim 7
Dependent Claim
58 words
A kit for simultaneously lysing cells and hybridizing released endogenous nucleic acid, comprising at least one vial containing a composition comprising: a buffer solution; at least one monomeric lactam at a concentration of about 5% to about 70% of the total volume of the buffer solution; and a detergent at a concentration of about 0.05% to about 0.5%.