Ideal primers for the PCR reaction should have the following feature: they should have a high A-T content the forward primer should be palindromic to the reverse primer they should anneal rapidly only on one target in the DNA template they should have a high G-C content

Ideal primers for the PCR reaction should have the following feature: they should have a high A-T content the forward primer should be palindromic to the reverse primer they should anneal rapidly only on one target in the DNA template they should have a high G-C content

Biochemistry

9th Edition

ISBN:9781319114671

Author:Lubert Stryer, Jeremy M. Berg, John L. Tymoczko, Gregory J. Gatto Jr.

Publisher:Lubert Stryer, Jeremy M. Berg, John L. Tymoczko, Gregory J. Gatto Jr.

Chapter1: Biochemistry: An Evolving Science

Section: Chapter Questions

Problem 1P

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Question

Ideal primers for the PCR reaction should have the following feature:

they should have a high A-T content
the forward primer should be palindromic to the reverse primer
they should anneal rapidly only on one target in the DNA template
they should have a high G-C content

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This is from the Wilson et al. article.
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Primers can sometimes bind and target the wrong gene, especially if the primers are allowed to bind to the DNA strands at a low temperature. PCR also preferentially amplify short segments of DNA. Would it be important to actually run the cDNA after the PCR on a DNA gel in order to check for a PCR product of the predicted size for the insulin gene? Why or why not? How many more genes in the plasmid (besides the insulin cDNA insert) are need to determine which bacterial cells have been transformed and to determine which transformed bacterial cells have a plasmid with the cDNA insulin insert? What is needed in the plasmid with the cDNA insert besides the gene for insulin to actually cause the bacteria to express the insulin gene and produce the insulin protein?
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