Question

Design PCR primers to amplify Kringles 2-4 of human plasminogen for directional subcloning into PBluescript. Show the exact sequence where the PCR product is directionally ligated (2 restriction enzymes) into the vector ( don't show all the intervening DNA of the PCR product)

Answer 1

Answer to the question #46782, Biology, Molecular Biology

Design PCR primers to amplify Kringles 2-4 of human plasminogen for directional subcloning into PBluescript. Show the exact sequence where the PCR product is directionally ligated (2 restriction enzymes) into the vector ( don't show all the intervening DNA of the PCR product.

Answer:

For (EcorI)

5'-GAATTCtgcgacattcttgagtgt-3'

Rev (BamHI)

5'-GGATTCcaacactcgcttctgttc-3'

Here is sequence of PCR product inserted into the vector. «|» marks place of ligation. Sequence of the vector is shown in blue, of the insert — in black.

-pBluescript--G|GATTCtgcgacattcttgagtgt---gaacagaagcgagtgttgCCTAG|G-pBluescript-pBluescript--CCTAA|Gacgctgtaagaactcaca---cttgtcttcgctcacaacG|GATCC-pBluescript-

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