Question

Regarding the trp operon leader sequence:

1.What would happen if the leader sequence was mutated in region 3, such that the it was missing?

2. Regarding Electrophoresis:

Restriction endonuclease (why restricted?, why endo?   Why -nucl-? Why - ase?)

                                                                                                                                                            

RFLP

Ethidium Bromide

Probe

3. DNA is ___________________ charged and therefore moves toward the _______________ electrode.

Answer 1

Q1

Tryptophan operon switch on when the concentration of tryptophan is low in the medium. This operon is regulated by leader sequence present upstream to the structural genes of the operon. Leader sequences has the four distinct regions. Presence of low concentration of tryptophan allows the formation of antitermination loop between sequence 2 and 3 of the leader sequence. Presence of high tryptophan leads to formation of termination loop between sequence 4 and 3. Therefore deleting sequence 3 would prevent the formation of termination loop and hence operon will be on even after presence of tryptophan. The formation of antitermination loop is not essential for the expression of tryptophan structural genes.

Q2

They are called as restricted because they cleaves the specific sequence of the DNA. That means cleavage is sequence specific. They are called as endo because they creat a cleavage within ( in between) the sequence of DNA. Nucl because they cleaves the nucleic acid. DNA is nucleic acid. Suffix "ase" added to the enzymes which catalyzed particular reaction. Restrictions endonucleases are the enzymes hence suffix "ase" is added.

RFLP is the restriction fragments length polymorphism. It is the technique used to know the variations in the homologous DNA sequences. In RFLP, presence of mutation in the one of the DNA fragment changes the restriction cleavage site and hence the different pattern of the DNA appeared upon gel electrophoresis.

Ethidium Bromide is added in the agarose gel to view the DNA. This reagent intercalated in between the DNA bases and gives coloration upon exposure to the ultraviolet light.

Probe is the labeled DNA sequence Used in the RFLP that hybridize with the digested DNA sequence after electrophoresis separation.

Q3

Negatively charged and moves to Anode (positivity charged) electrode.

Explanation.

Presence of phosphate gives negative charge to the DNA molecule. Therefore, it moves to the opposite charge which is positively charged anode electrode.