Method of amplifying DNA and RNA mismatch cleavage products

Details Method of amplifying DNA and RNA mismatch cleavage products Method of amplifying DNA and RNA mismatch cleavage products

1998-09-10

2002-09-24

Campbell, Eggerton A. (Department: 1656)

Chemistry: molecular biology and microbiology

Measuring or testing process involving enzymes or...

Involving nucleic acid

C536S023100

Reexamination Certificate

active

06455249

ABSTRACT:

FIELD OF THE INVENTION
The present invention generally relates to detection of nucleic acid sequence mutations. The present invention more specifically relates to a new and useful method for signal amplification of mismatch cleavage.
BACKGROUND OF THE INVENTION
The ability to detect alterations in nucleic acid sequences (for example, mutations and polymorphisms) is central to the diagnosis of genetic diseases and to the identification of clinically significant variants of disease-causing microorganisms. Similarly, identification and measurement of RNA is necessary for determining control of gene transcription.
One method for the molecular analysis of genetic variation involves the detection of restriction fragment length polymorphisms (RFLPs) using the Southern blotting technique (Southern, E. M.,
J. Mol. Biol
., 98 503-517, 1975). Since this approach is relatively cumbersome, new methods have been developed, some of which are based on the polymerase chain reaction (PCR).
These include: RFLP analysis using PCR (Chehab et al.,
Nature
, 329, 293-294, 1987; Rommens et al.,
Am. J. Hum. Genet
., 46, 395-396, 1990), allele-specific amplification (ASA) (Newton C R et al.,
Nuc. Acids Res
., 17, 2503-2516, 1989), oligonucleotide ligation assay (OLA) (Landergren U et al.,
Science
241, 1077-1080, 1988), primer extension (Sokolov B P,
Nucl. Acids Res
., 18, 3671, 1989), artificial introduction of restriction sites (AIRS) (Cohen L B et al.,
Nature
334, 119-121, 1988), allele-specific oligonucleotide hybridization (ASO) (Wallace R B et al.,
Nucl. Acids Res
., 9, 879-895, 1981) and their variants.
The following are further examples of art discussing mismatch repair enzymes and systems utilizing such enzymes in addition to other related subject matter:
Lu et al., 80
Proc. Natl. Acad. Sci. USA
4639, 1983 disclose the use of a soluble
E. coli
system to support mismatch correction in vitro.
Pans et al., 163
J. Bact
. 1007, 1985 disclose cloning of the mutS and mutL genes of
Salmonella typhimurium.
The specific components of the
E. coli
mispair correction system have been isolated and the biochemical functions determined. Preparation of MutS protein substantially free of other proteins has been reported (Su and Modrich, 1986
, Proc. Nat. Acad. Sci. U.S.A
., 84, 5057-5061. The isolated MutS protein was shown to recognize four of the eight possible mismatched base pairs (specifically, G-T, A-C, A-G and C-T mispairs).
U.S. Pat. No. 5,556,750 (“Methods and kits for fractionating a population of DNA molecules based on the presence or absence of a base-pair mismatch utilizing mismatch repair systems, issued Sep. 17, 1996 to Duke University) describes methods of fractionating DNA molecules based on base-pair mismatch utilizing mismatch repair systems.
Su et al., 263
J. Biol. Chem
. 6829, 1988 disclose that the mutS gene product binds to each of the eight base pair mismatches and does so with differential efficiency.
Jiricny et al., 16
Nucleic Acids Research
7843, 1988 disclose binding of the mutS gene product of
E. coli
to synthetic DNA duplexes containing mismatches to correlate recognition of mispairs and efficiency of correction in vivo. Nitrocellulose filter binding assays and band-shift assays were utilized.
Welsh et al., 262
J. Biol. Chem
. 15624, 1987 purified the product of the MutH gene to near homogeneity and demonstrated the MutH gene product to be responsible for d(GATC) site recognition and to possess a latent endonuclease that incises the unmethylated strand of hemimethylated DNA 5′ to the G of d(GATC) sequences.
Au et al., 267
J. Biol. Chem
. 12142, 1992 indicate that activation of the MutH endonuclease requires MutS, MutL and ATP.
Grilley et al. 264
J. Biol. Chem
. 1000, 1989 purified the
E. coli
mutL gene product to near homogeneity and indicate that the mutL gene product interacts with MutS heteroduplex DNA complex.
Lahue et al., 245
Science
160, 1989 delineate the components of the
E. coli
methyl-directed mismatch repair system that function in vitro to correct seven of the eight possible base pair mismatches. Such a reconstituted system consists of MutH, MutL, and MutS proteins, DNA helicase II, single-strand DNA binding protein, DNA polymerase III holoenzyme, exonuclease I, DNA ligase, ATP, and the four deoxyribonucleoside triphosphates.
Su et al., 31
Genome
104, 1989 indicate that under conditions of restricted DNA synthesis, or limiting concentration of dNTPs, or by supplementing a reaction with a ddNTP, there is the formation of excision tracts consisting of single-stranded gaps in the region of the molecule containing a mismatch and a d(GATC) site.
Grilley et al. 268
J. Biol. Chem
. 11830, 1993, indicate that excision tracts span the shorter distance between a mismatch and the d(GATC) site, indicating a bidirectional capacity of the methyl-directed system.
Holmes et al., 87
Proc. Natl. Acad. Sci. USA
, 5837, 1990, disclose nuclear extracts derived from HeLa and Drosophila melanogaster K[c] cell lines to support strand mismatch correction in vitro.
Cooper et al., 268
J. Biol. Chem
., 11823, 1993, describe a role for RecJ and Exonuclease VII as a 5′ to 3′ exonuclease in a mismatch repair reaction. In reconstituted systems such a 5′ to 3′ exonuclease function had been provided by certain preparations of DNA polymerase III holoenzyme.
Au et al., 86
Proc. Natl. Acad. Sci. USA
8877, 1989 describe purification of the mutY gene product of
E. coli
to near homogeneity, and state that the MutY protein is a DNA glycosylase that hydrolyzes the glycosyl bond linking a mispaired adenine (G-A) to deoxyribose. However, their enzyme did not cleave the A strand at “A” in a circular closed heteroduplex DNA with G/A mismatch as a substrate. The MutY protein, DNA polymerase I, and DNA ligase were shown to reconstitute G-A to G-C mismatch correction in vitro in the presence of an apurinic endonuclease.
Tsai-Wu et al., 89
Biochemistry
8779, 1992, cloned mutY gene, overexpress and purified the MutY enzyme to homogeneity for examining enzyme specificity. In addition to glycosylase activity, this MutY enzyme can cleave the “A” of G/A mismatch on the A strand.
Wiebauer and Jiricny, 339
Nature
234, 1989, discovered the correction of G/T mispairs to G/C pairs by thymine DNA glycosylase in human cells.
Nedderman and Jiricny, 268
J. Biol. Chem
. 21218, 1993, purified the G/T mnispair specific thymine glycosylase from HeLa cells.
Slupsker et al., 178
, J. Bacteriol
. 3885, 1996, cloned and sequenced a human homolog (hMYH) of
E. coli
mutY gene whose function is the repair of oxidative DNA damage.
A role for the
E. coli
mismatch repair system in controlling recombination between related but non-allelic sequences has been indicated (Feinstein and Low, 113
Genetics
, 13, 1986; Rayssiguier, 342
Nature
396, 1989; Shen, 218
Mol. Gen. Genetics
358, 1989; Petit, 129
Genetics
327, 1991). The frequency of crossovers between sequences which differ by a few percent or more at the base pair level are rare. In bacterial mutants deficient in methyl-directed mismatch repair, the frequency of such events increases dramatically. The largest increases are observed in MutS and MutL deficient strains. (Rayssiguier, supra; and Petit, supra.)
Nelson et al., 4
Nature Genetics
11, 1993, disclose a genomic mismatch (GMS) method for genetic linkage analysis. The method allows DNA fragments from regions of identity-by-descent between two relatives to be isolated based on their ability to form mismatch-free hybrid molecules.
During the last few years, a group of bacteria repair enzymes, e.g., endo VIII (Melamede, et al.,
Biochemistry
, 33, 1255-1264, 1994), endo III (Dizdaroglu et al.,
Biochemistry
, 32, 12105-12111, 1993), formamidopyrimidine-DNA glycosylase (Chetsanga et al.,
Biochemistry
, 20, 5201-5207, 1981), T4 endonuclease V (Nakabeppu et al.,
J. Biol. Chem
., 257, 2556-2562, 1982; McCullough et al.,
J. Biol. Chem
., 272, 27210-27217, 1997), etc., have been isolated and characterized to repair the damaged and modified nucleic aci

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