In this question, determine the values for the linking number (L), super helical

Question

In this question, determine the values for the linking number (L), super helical turns (AL), the twisting number (T) and the writhing number (W supercoiling number) initially and after Steps 1-4. Each part of your answer will be for the plasmid as a whole. It will be helpful to keep track of whether a nick is present or not. Reference Init Step Step2 Step 3 Step 4 Linking (L) 1.inking (AL) Twisting (T) Writhing (W) Supercoiling) NICK? You decide No YES YES NO No Below the table, explain your answers. 1.Choose a reference state in the way suggested in class. Once you have chosen this reference state at the outset of the problem, do not change it at any time throughout the problem. (After all, what good is a reference state if it is not constant) 2. Throughout this problem, assume that all duplex DNA is B-DNA (I0 bp/turn), except if it is mentioned otherwise. Initial State: A 5000 bp plasmid is completely in the B-DNA conformation under normal conditions and has 25 underwindings Step I: The plasmid is nicked once in one strand and continues to be B-DNA Step 2: There is a region of 240 bp in the plasmid that is normally in the B-DNA conformation, but can be induced to adopt the Z-DNA conformation by changing the salt concentration. The plasmid DNA is manipulated such that this B-DNA to Z-DNA transition occurs in the 240bp region. The rest of the plasmid remains B-DNA Important Hints: .After step 2, the plasmid is "relaxed", since it remains nicked. HOWEVER, it is not in the same relaxed state as the reference state. In spite of the change in salt concentration, do not change your reference state 2. Some information about Z-DNA (not all of which may be pertinent): Z-DNA has 12 bp/turn: Z- DNA is left-handed; in Z-DNA the sugars altenate between 2- and 3-endo: the bases alternate between syn- and anti-conformation Step 3: The plasmid is re-ligated (i.e, the nick is re-sealed) Step 4The plasmid is returned to the original salt concentration, and all of the plasmid regains the original B-DNA conformation. (Note: no nicking occurred in Step 4.) In summary the manipulations and questions can be outlined as follows: Initial Step 1 Step 2 Step 3 religate Step 4 startingnicked change 240bp plasmid change 240bp plasmid region to 2-DNA region back to B-DNA

Explanation / Answer

Answer

The linking number defines the number of times a strand of DNA winds in the right-handed direction around the helix axis.

circular DNA of 5000 bp,

the linking number

=5000/10

= 500,

10 is the basepair per turn for type B DNA

form B type helix the preferred twist = number of base pair/10;

10 is the number of the nucleotide in one twist.

= 5000/ 10

= 500

A right-handed coil is assigned a negative number and the left-handed coil is assigned a positive number.

Initial state L = 500, delta L = 0, T = 500, W = 25

Step 1: L= 500, delta L = 0, T = 500, W = 25

Step 2: L= 500, T= 0, W= 0

Step 3: L= 500, T= 500, W= 0

Step 4: L= 500, T= 500, W= 25

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