Question

4. Imagine you cloned a PCR product (1500 bp) into a plasmid that is 3000 bp long. You know that the enzyme BamHI has just one site in the plasmid very close to the PCR product. You know that the enzyme Xmal has two sites in the plasmid, both very close to each end of the PCR product in the plasmid. (5 points) You set up some restriction digests to analyze your cloning: 1. Uncut plasmid control 2. BamHI plasmid digestion 3. Xmal plasmid digestion Draw the results you expect to see on the gel, if your cloning and digests worked exactly as expected. Ladder Kilobases 10.0 8.0- 6.0 5.0 4.0 3.0 2.0- 1.5 1.2 1.0 0.9 0.8 0.7 0.6 0.5 0.4 0.3 0.2 Uncut BamHI Xmal 0.1

          4. Imagine you cloned a PCR product (1500 bp) into a plasmid that is 3000 bp long. You
know that the enzyme BamHI has just one site in the plasmid very close to the PCR product.
You know that the enzyme Xmal has two sites in the plasmid, both very close to each end
of the PCR product in the plasmid. (5 points)
You set up some restriction digests to
analyze your cloning:
1. Uncut plasmid control
2. BamHI plasmid digestion
3. Xmal plasmid digestion
Draw the results you expect to see on
the gel, if your cloning and digests
worked exactly as expected.
Ladder
Kilobases
10.0
8.0-
6.0
5.0
4.0
3.0
2.0-
1.5
1.2
1.0
0.9
0.8
0.7
0.6
0.5
0.4
0.3
0.2
Uncut
BamHI
Xmal
0.1

Show more…

4. Imagine you cloned a PCR product (1500 bp) into a plasmid that is 3000 bp long. You
know that the enzyme BamHI has just one site in the plasmid very close to the PCR product.
You know that the enzyme Xmal has two sites in the plasmid, both very close to each end
of the PCR product in the plasmid. (5 points)
You set up some restriction digests to
analyze your cloning:
1. Uncut plasmid control
2. BamHI plasmid digestion
3. Xmal plasmid digestion
Draw the results you expect to see on
the gel, if your cloning and digests
worked exactly as expected.
Ladder
Kilobases
10.0
8.0-
6.0
5.0
4.0
3.0
2.0-
1.5
1.2
1.0
0.9
0.8
0.7
0.6
0.5
0.4
0.3
0.2
Uncut
BamHI
Xmal
0.1

Added by Michael C.

Biology for AP Courses

Julianne Zedalis, John Eggebrecht

Instant Answer

Solved by Expert Jenny Wu

06/07/2023

Step 1

The plasmid is 3000 bp long and has one BamHI site and two Xmal sites. The PCR product is 1500 bp long. Show more…

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Texts: 4. Imagine you cloned a PCR product (1500 bp) into a plasmid that is 3000 bp long. You know that the enzyme BamHI has just one site in the plasmid very close to the PCR product. You know that the enzyme Xmal has two sites in the plasmid, both very close to each end of the PCR product in the plasmid. (5 points) You set up some restriction digests to analyze your cloning: Ladder Uncut BamHI Xmal Kilobases 10.0 8.8 3.0 2.0 Draw the results you expect to see on the gel, if your cloning and digests worked exactly as expected. 1.5 1.2 1.0 0.9 0.8 0.7 0.6 0.5 0.4 0.3 0.2 0.1

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Transcript

00:01 For one, you want to clone a fragment of dna into a plasmid vector such as pbr322.

00:06 The plasmid is 4 .36 kv long and has a single restriction site for the restriction and then bamh1.

00:13 Now, if you digest the pbr322 with bamh1, how many linear fragments would you have? so this is pbr322.

00:27 It has a single bamh1 site.

00:37 So let's say it's here.

00:38 So once you cut the plasmid with the bamh1, the enzyme is going to recognize the bamh1 site and cut it.

00:50 So it only is going to cut once right here.

00:54 So once you cut the circular plasmid once with bamh1 enzyme, it is going to linearize.

01:04 And the size of this fragment is going to be 4 .36 kv.

01:10 And you only produce one fragment.

01:13 So the correct answer is going to be a, one.

01:16 And this fragment is 4 .36 kv long.

01:21 So basically you cut a circle open so it become linearized.

01:26 So this is the first question.

01:28 So i draw the figure to show you what it looks like.

01:31 The second question.

01:33 Now, you have pbr322 has a single bamh1 site and a single bspmi site.

01:41 A double digestion produced two fragment of approximately 690 base pairs, 3670 base pair.

01:49 Map the two sites on a circular plasmid.

01:53 If the bamh1 is at the beginning of the plasmid, tetracycline resistant gene, and a gene is about 1200 base pair long, is the bamh1, bspmi site also in the gene? so let's say you have a plasmid circular...

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