Introduction to Bioinformatics II Restriction enzyme Lecture 15 By Ms. Shumaila Azam

Restriction Endonuclease Proteins that recognize particular sequence of nucleotide (called the restriction site and generally 4 to 8 bases long) and cut the double stranded DNA molecule at restriction site

Classification Three classes Differ in their structure They cut the DNA substrate at their recognition site The recognition and the cut sites are separate from each other.

Restriction enzymes Its recognition site has a palindromic sequence E.g GTAATG (on a single strand) GTATAC (on double strand) CATATG To cut DNA , all the restriction enzymes make two incisions, once through each sugar-phosphate backbone of the DNA. More than 3000 enzymes are studied More than 600 enzymes are available commercially

RFLP Restriction Fragment Length Polymorphism (RFLP) Widely used laboratory method in molecular identification and Phylogenetic studies. This approach requires the sequences to be cut into several fragments with the help of restriction endonucleases. The variation in the position of these sites along the DNA, among the sequences being analyzed will lead to digested product that are of varying lengths. Following a high-resolution gel electrophoresis of the digested product, the fragment-patterns are visually compared to determine the similarity between the sequences.

RFLP

Proposed Concept Enzyme Cut Order (ECO) – information from DNA for evaluation Definition: ECO for a DNA sequence (S) for a particular set of restriction enzymes {Ez} is a string (array) of enzyme names (represented as numeric id) in the order each enzyme (ez Є Ez) cuts the sequence. ECO may also include position of nucleotide from the start of sequence where the cut occur. Thus, ECO is a string (array) of tuples consisting of enzyme id and cut position. Example:

GenBank Classification

Concept Contd.. Closely related organisms have similar Enzyme Cut Order Table1 : The ECO for ‘ITS’ sequences from close and distantly related fungi. The closely related Nectria species (Nectria haematococca and Nectria mauritiicola) show high level of ECO similarity.

ECO Enzyme Cut Order (ECO)- Similarity Score The similarity score between two ECO consists Number of similar enzymes and Order in which these enzyme cut the sequence The similarity score will be higher if we find larger number of similar enzymes appearing in the same order among two Enzyme Cut Orders. This similarity score is the Longest Common Subsequence (LCS) among two strings – the strings are the ECO The length of Longest Common Subsequence (LCS) between two ECO (E1 and E2) of two corresponding sequences (S1 and S2) are considered as the Enzyme Cut Order Similarity Score between E1 and E2.

Result Summary Enzyme Cut Order is a distinguishing characteristic of DNA sequences The similarity between two sequences can be defined by Enzyme Cut Order Similarity Score ECO-similarity score can be measured as the length of LCS among the corresponding Enzyme Cut Orders of the DNA sequences of the organisms