Restriction Enzymes Flashcards
What are restriction enzymes (restriction endonuclease)?
They are a special class of DNA cutting enzymes that exist in many bacteria and also known as molecular scissors.
What’s the importance of restriction enzymes for bacteria?
The restriction enzyme prevents replication of the phage DNA by cutting it into many pieces.
Why are restriction enzymes called by this name?
Due to their ability to restrict, or limit, the number of bacteriophage that can infect a bacterium.
What are recognition sequences?
They are very short, specific sequence of nucleotide bases (4 to 6 bases) which are randomly distributed throughout the DNA.
How does bacteria disguise their recognition sequences? and why?
Enzymes called methylases add methyl groups (-CH3) to adenine or cytosine bases within the recognition sequence, which is thus modified and protected from the endonuclease. They do that in order to prevent their DNA from being degraded.
Why are recognition sequences usually palindromic sequences?
Because their recognition sequence reads the same in the 5’ to 3’ direction on both DNA strands.
What are the different types of restriction enzymes?
There are four different categories of restriction enzymes :
1) Type I
2) Type II
3) Type III
4) Type IV
Which type of restriction enzymes the most useful for laboratory experiments? and why?
Because they cut within or close to their recognition sequence.
What is DNA cloning or recombinant DNA technology?
Type II restriction enzymes are used to cut DNA molecules at interesting specific locations and then reattach different DNA sequences to each other using an enzyme called DNA ligase, creating new, recombined DNA sequences, or essentially new DNA molecules.
What are sticky ends?
They are stretches of single-stranded DNA at the ends of the DNA fragments. They join spontaneously by hydrogen boding (base pairing) with complementary overhangs..
What are blunt ends?
They are produced when both strands of DNA are cut in the same place with no complementary overhangs.
Mention different restriction enzymes and what ends they produce.
1) BamHI produces sticky ends.
2) EcoRI produces sticky ends.
3) HaeIII produces blunt ends.
4) EcoR V produces blunt ends.
5) HindIII produces sticky ends.
6) Pst I produces sticky ends.
7) Not I produces sticky ends.
What happens when two fragments of DNA from different sources have been produced by the same restriction enzyme (كده عندهم نفس ال recognition site)?
The two pieces will have identical sets of sticky ends and can be spliced (recombined) in vitro.
What does in vitro mean?
When something happens outside the organism.
What type of bond does DNA ligase and sticky ends sticking together produce?
1) DNA ligase covalently (covalent bond) link the backbones of the DNA pieces.
2) Sticky ends sticking together by hydrogen bonds ( base pairing ).
