You ran your uncut plasmid DNA on a gel in Molecu lar Biology (Part II). Downloa

Question

You ran your uncut plasmid DNA on a gel in Molecu lar Biology (Part II). Download your gel picture from the Molecular- III folder for your instructor/lab section on BOX and interpret your gel results to answer the following: A. Do you have plasmid DNA to work with? How do you know? B. If your answer to 1A is 'yes', can you tell if your plasmid DNA is of the expected size? Explain. If your answer tolA is 'no', explain what you think happened. *Even if you were unsuccessful at obtaining plasmid DNA in the miniprep, you must still complete the remainder of the assignment to design a restriction digest experiment. For your restriction digest experiment, what will you use as a control? What reagents will the control sample contain?

Explanation / Answer

1. A) yes, the image of gel electrophoresis cleary is showing 2 bands in sample section which represents the supercoiled and relaxed form of plasmid the first coloumn represents the ladder that is used as standard and also to deduce the size of the sample. there should be a third band representing the linear form of plasmid but as the plasmid sample was uncut it will not have its linear form.
?B) to known whether your plasmid size , we compare the bands obtained in sample column to the ladder used in electrophoresis. now among the two bands of plasmid we will use the higher one as it is the relaxed form of plasmid as it will give much closer image of its size. Generally the ladder used in such experiments for plasmid the its size ranges from 0.5 kb to 10kb /1Mb in both the cases the plasmid we obtaind is of expected size as the expected size of a plasmid in its relaxed state is from 2kb to 1Mb

2. the control is that sample which remains untreated with experiment conditions. i.e control sample will only contain untreated sample plus buffer but no reagents to ensure that whatever results that we will obtained are becuase of the reagents / condition we used to treat that intial sample. we keep every condition same except one condition whose difference we are trying to test.
?for restriction digestion of plasmid we want to understand the effect of restriction enzyme on plasmid therefore we treat our plasmid with restriction enzymes and then pour them into the agrose gel and the control used in this experiment is the untreated plasmid ?and this sample will contain everything same as a treated sample except the enzyme,(i.e. buffer +water +untreated plasmid DNA+ tracking dye ) ?by doing this we get to know the actual plasmid DNA integrity.
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