4) An unknown plasmid (circular DNA) is digested with Eco RI...

4) An unknown plasmid (circular DNA) is digested with Eco RI and Bam HI, and the fragments are separated by a gel electrophoresis. A diagram of the resulting gel is shown. Use this information to create a restriction map. Eco RI kb Eco RI Bam HI Bam HI kb 3.0 3.0 2.2 --- 1.9 --- --- 1.9 1.4 --- --- 1.5 --- --- 1.2 --- 1.1 --- 1.0 --- 0.3 Hint: Add up the fragments from each column to determine the total length of the original DNA. Also you may want to draw a map of possible Eco RI cuts and Bam HI cuts separately first. This should help you visualize where the cuts occur when both enzymes are present.

Transcript

00:01 All right, so based on what we have seen in the electrophoresis gel, this is now our restriction map.

00:10 Okay, so let's go ahead and do the products first.

00:13 Let's confirm by looking at the sizes, right? so first we need to, let's show the ecor1 cut sides, right? so we have a 1 .4 kb product here.

00:29 That's not very circular, is it? let's try again.

00:38 Oh, a little bit better.

00:39 Okay, so we have a 1 .4 fragment here, a 2 .2 kb fragment there, and then the 1 .9 fragment here.

00:56 This 1 .9 fragment is going to be really important because that basically just means that that's the one that has no bam h1 cut site.

01:05 I say this because if you look at the double digest, that is the one band that is not, that is present in complete size.

01:17 There's also for ecor1 there's also a 1kb size fragment that is present in bamh1 that is also present in the double digest but if you take a look those other fragments will actually if you add up some of the fragments that will equal the sizes of the ecor1 cut sites the one thing that we cannot add the other fragment sizes up to is 1 .9kb since that is the case that 1 .9 kb fragment is not it means that that means that the 1 .9 kb fragment has no bm h1 cut site okay so basically um we have this here now let's look at the the uh bam h1 fragments so we have a 3kb size uh which is going to be on this end right here so that's going to be well let's do it better this is in total going to be 3kb then we have 1 .5 and 1 .5 and 1 1 .5 and 1 kb fragments, okay? so, where does that lead us with the double digests? let's use green.

03:16 All right, so if you look at the double digest, we have our 1 .9kb fragment, which is here.

03:22 That might be too dark.

03:23 Let's make it lighter...

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