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Isocaudomers are pairs of restriction enzymes that have slightly different recognition sequences, but upon cleavage of DNA, generate identical overhanging termini sequences. These sequences can be ligated to one another, but then form an asymmetrical sequence that cannot be cleaved by a restriction enzyme.
For example the enzymes Mbo I and BamH I are isocaudomers:
Mbo I N*GATC N N CTAG*N BamH I G*GATC C C CTAG*G
N represents any of the four nucleotides. Independently of which nucleotide is present when cleaving with MboI, after cleavage with either enzyme, all termini have the central tetranucleotide - GATC. This allows fragments generated with one enzyme to anneal with fragments generated with the other enzyme. This can be used for elimination of restriction sites from the resulting DNA fragment. For example:
Not I GC*GGCC GCCG CCGG*CG Bsp120 I G*GGCC CC CCGG*G
In the above example, both enzymes produce tetranucleotides CCGG which can anneal to one another. However, resulting DNA sequence will be:
GCGGCCCCGCCGGG
where the nucleotides shown in italic originate from NotI-cut site, and those in bold from Bsp120I-cut one. Note that the resulting sequence is not recognised by either of the two enzymes.
Other examples of isocaudomers [1] include:
BamHI/BclI/BglII/BstYI/DpnII
NcoI/BspHI/FatI/PciI
NdeI/AseI/BfaI/Csp6I/MseI
XbaI/AvrII/NheI/SpeI/StyI
XhoI/PspXI/SalI
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A restriction enzyme, restriction endonuclease, REase, ENase orrestrictase is an enzyme that cleaves DNA into fragments at or near specific recognition sites within molecules known as restriction sites. Restriction enzymes are one class of the broader endonuclease group of enzymes. Restriction enzymes are commonly classified into five types, which differ in their structure and whether they cut their DNA substrate at their recognition site, or if the recognition and cleavage sites are separate from one another. To cut DNA, all restriction enzymes make two incisions, once through each sugar-phosphate backbone of the DNA double helix.
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In biochemistry, a nuclease is an enzyme capable of cleaving the phosphodiester bonds between nucleotides of nucleic acids. Nucleases variously effect single and double stranded breaks in their target molecules. In living organisms, they are essential machinery for many aspects of DNA repair. Defects in certain nucleases can cause genetic instability or immunodeficiency. Nucleases are also extensively used in molecular cloning.
A cDNA library is a combination of cloned cDNA fragments inserted into a collection of host cells, which constitute some portion of the transcriptome of the organism and are stored as a "library". cDNA is produced from fully transcribed mRNA found in the nucleus and therefore contains only the expressed genes of an organism. Similarly, tissue-specific cDNA libraries can be produced. In eukaryotic cells the mature mRNA is already spliced, hence the cDNA produced lacks introns and can be readily expressed in a bacterial cell. While information in cDNA libraries is a powerful and useful tool since gene products are easily identified, the libraries lack information about enhancers, introns, and other regulatory elements found in a genomic DNA library.
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A restriction digest is a procedure used in molecular biology to prepare DNA for analysis or other processing. It is sometimes termed DNA fragmentation, though this term is used for other procedures as well. In a restriction digest, DNA molecules are cleaved at specific restriction sites of 4-12 nucleotides in length by use of restriction enzymes which recognize these sequences.
Mung bean nuclease is a nuclease derived from sprouts of the mung bean that removes nucleotides in a step-wise manner from single-stranded DNA molecules (ssDNA) and is used in biotechnological applications to remove such ssDNA from a mixture also containing double-stranded DNA (dsDNA). This enzyme is useful for transcript mapping, removal of single-stranded regions in DNA hybrids or single-stranded overhangs produced by restriction enzymes, etc. It has an activity similar to Nuclease S1, but it has higher specificity for single-stranded molecules.
HindIII (pronounced "Hin D Three") is a type II site-specific deoxyribonuclease restriction enzyme isolated from Haemophilus influenzae that cleaves the DNA palindromic sequence AAGCTT in the presence of the cofactor Mg2+ via hydrolysis.
HaeIII is one of many restriction enzymes (endonucleases) a type of prokaryotic DNA that protects organisms from unknown, foreign DNA. It is a restriction enzyme used in molecular biology laboratories. It was the third endonuclease to be isolated from the Haemophilus aegyptius bacteria. The enzyme's recognition site—the place where it cuts DNA molecules—is the GGCC nucleotide sequence which means it cleaves DNA at the site 5′-GG/CC-3. The recognition site is usually around 4-8 bps.This enzyme's gene has been sequenced and cloned. This is done to make DNA fragments in blunt ends. HaeIII is not effective for single stranded DNA cleavage.
BamHI is a type II restriction endonuclease, having the capacity for recognizing short sequences of DNA and specifically cleaving them at a target site. This exhibit focuses on the structure-function relations of BamHI as described by Newman, et al. (1995). BamHI binds at the recognition sequence 5'-GGATCC-3', and cleaves these sequences just after the 5'-guanine on each strand. This cleavage results in sticky ends which are 4 bp long. In its unbound form, BamHI displays a central b sheet, which resides in between α-helices.
Deoxyribonuclease IV (phage-T4-induced) is catalyzes the degradation nucleotides in DsDNA by attacking the 5'-terminal end.
BglII is a type II restriction endonuclease isolated from certain strains of Bacillus globigii.
PstI is a type II restriction endonuclease isolated from the Gram negative species, Providencia stuartii.
A primer dimer (PD) is a potential by-product in the polymerase chain reaction (PCR), a common biotechnological method. As its name implies, a PD consists of two primer molecules that have attached (hybridized) to each other because of strings of complementary bases in the primers. As a result, the DNA polymerase amplifies the PD, leading to competition for PCR reagents, thus potentially inhibiting amplification of the DNA sequence targeted for PCR amplification. In quantitative PCR, PDs may interfere with accurate quantification.
In molecular biology, XhoI is a type II restriction enzyme EC that recognise the double-stranded DNA sequence CTCGAG and cleaves after C-1. Type II restriction endonucleases are components of prokaryotic DNA restriction-modification mechanisms that protect the organism against invading foreign DNA. These site-specific deoxyribonucleases catalyse the endonucleolytic cleavage of DNA to give specific double-stranded fragments with terminal 5'-phosphates.
Arthrobacter luteus (ALU) is a species of gram-positive bacteria in the genus Arthrobacter. A. luteus is facultatively anaerobic, pleomorphic, branching, non-motile, non-sporulating, non-acid-fast, catalase-positive, and rod-shaped.
EcoRI is a restriction endonuclease enzyme isolated from species E. coli. It is a restriction enzyme that cleaves DNA double helices into fragments at specific sites, and is also a part of the restriction modification system. The Eco part of the enzyme's name originates from the species from which it was isolated - "E" denotes generic name which is "Escherichia" and "co" denotes species name, "coli" - while the R represents the particular strain, in this case RY13, and the I denotes that it was the first enzyme isolated from this strain.
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