Biomaterial Database

S1 nuclease does not cleave DNA at single-base mis-matches. 1981

J R Silber, and L A Loeb

Three assays have been designed to detect the cleavage of duplex phi X174 DNA at single-base mis-matches. Studies with S1 nuclease failed to detect cleavage at mis-matches. S1 nuclease digestion at 37 and 55 degrees C failed to produce a preferential degradation of a multiply mis-matched heteroduplex when compared to a mis-match-free homo-duplex as analyzed by sedimentation on sucrose gradients. Other heteroduplex templates were not cleaved by S1 nuclease at a defined single-base mis-match when assayed by gel electrophoresis or by marker rescue. In all cases, the amount of S1 nuclease employed was at least 10-times more than that required to render a single-stranded phi X174 DNA molecule completely acid soluble. The rate of hydrolysis of single-base mis-matches by S1 nuclease was estimated to be less than 0.016% of the rate at a base in single-strand phi X174 DNA. In no instance did we detect activity by S1 nuclease directed at mis-matched sites in our template molecules. Similarly, the single-strand specific endonuclease from Neurospora crassa does not cleave heteroduplex templates at a defined single-base mis-match when assayed by marker rescue.

UI	MeSH Term	Description	Entries
D010584	Bacteriophage phi X 174	The type species of the genus MICROVIRUS. A prototype of the small virulent DNA coliphages, it is composed of a single strand of supercoiled circular DNA, which on infection, is converted to a double-stranded replicative form by a host enzyme.	Coliphage phi X 174,Enterobacteria phage phi X 174,Phage phi X 174,phi X 174 Phage,Phage phi X174
D002499	Centrifugation, Density Gradient	Separation of particles according to density by employing a gradient of varying densities. At equilibrium each particle settles in the gradient at a point equal to its density. (McGraw-Hill Dictionary of Scientific and Technical Terms, 4th ed)	Centrifugations, Density Gradient,Density Gradient Centrifugation,Density Gradient Centrifugations,Gradient Centrifugation, Density,Gradient Centrifugations, Density
D004279	DNA, Viral	Deoxyribonucleic acid that makes up the genetic material of viruses.	Viral DNA
D004720	Endonucleases	Enzymes that catalyze the hydrolysis of the internal bonds and thereby the formation of polynucleotides or oligonucleotides from ribo- or deoxyribonucleotide chains. EC 3.1.-.	Endonuclease
D006868	Hydrolysis	The process of cleaving a chemical compound by the addition of a molecule of water.
D001483	Base Sequence	The sequence of PURINES and PYRIMIDINES in nucleic acids and polynucleotides. It is also called nucleotide sequence.	DNA Sequence,Nucleotide Sequence,RNA Sequence,DNA Sequences,Base Sequences,Nucleotide Sequences,RNA Sequences,Sequence, Base,Sequence, DNA,Sequence, Nucleotide,Sequence, RNA,Sequences, Base,Sequences, DNA,Sequences, Nucleotide,Sequences, RNA
D013379	Substrate Specificity	A characteristic feature of enzyme activity in relation to the kind of substrate on which the enzyme or catalytic molecule reacts.	Specificities, Substrate,Specificity, Substrate,Substrate Specificities
D015719	Single-Strand Specific DNA and RNA Endonucleases	Enzymes that catalyze the endonucleolytic cleavage of single-stranded regions of DNA or RNA molecules while leaving the double-stranded regions intact. They are particularly useful in the laboratory for producing "blunt-ended" DNA molecules from DNA with single-stranded ends and for sensitive GENETIC TECHNIQUES such as NUCLEASE PROTECTION ASSAYS that involve the detection of single-stranded DNA and RNA.	Single Strand Specific DNA and RNA Endonucleases