Attached

1) Marking and labeling the following gels with the molecular weight marker bands as follows:
a) We use molecular weight marker bands in gel of different known size to estimate the size of an unknown macromolecule such as DNA or protein. Based on the electrical mobility and size of the molecules, they get separated in the polyacrylamide or agarose gel which allows us to use them as reference for estimation.
Pfu polymerase is a protein of molecular weight 90 kDa. Thus, in the gel A, lanes 4-10 show the correct size of the polymerase which is marked in the above figure in reference to the molecular weight marker bands available.
b) Lane 2 represents lysate from E. coli BL21 (DE3) without any expression of the pET vectors for Pfu as well as for Pwo. They are the negative controls that indicate the proteins present naturally in the E. coli BL21 system, which can help us to detect the changes or variables in the experiment after the expression of the desired vector. They help us to ensure that the used experimental procedures are valid.
Lane 3 represents the positive control or lysate from E. coli BL21 with the expression of pETPfu (A) and pETPwo (B). They us to identify and check the expression of the desired protein in the lysate for purification. Also, they to validate the correct experimental procedure.
c) Lanes 2 and Lanes 3 represents the negative and positive control of the experiment. They are usually used to validate the experimental procedure such that to confirm they are not any placebo effect. The negative control showcases the naturally expressing protein and the positive control clearly shows the difference in the expressing protein after the expression of desired vector or treatment. Also, the positive control helps us to identify any problems in the experiment as the results of the lane are already benchmarked.
d) Lanes 4-10 represents the protein purification information. Lane 4 determines the cleared lysate along with Nickel NTA beads from the total lysate pool of E. coli BL21 after denaturing the protein with heat treatment. As the desired proteins are His tagged, they are mostly cleared or purified from the lysate using Nickel NTA beads using affinity chromatography. Lane 5 and 6...