You perform electrophoresis on the product of a PCR and see that instead of the

Question

You perform electrophoresis on the product of a PCR and see that instead of the 300-baseband you expected that there are five different bands.

What controls the specific sequence to which a PCR primer binds? complementary sequence

What are two factors that contribute to the annealing temperature of the primer? primer length and the GC content (What does GC stand for)

What part of your thermal cycle protocol could you change, and how would you change it, to try and get rid of the extra bands?

      Thermal cycle protocol could you change -

      Change it - temperature should be increased to try to get rid of the extra bands.

d. If, after performing electrophoresis, there were no bands on the gel, what part of your thermal cycle protocol would you change, and how would you change it?

Thermal cycle protocol could you change -

Change it - temperature should be decreased to allow binding of the primer.

If you are using a DNA sequence from a related species to design primers for your species of interest, why is it better to design your primers to complement an exon?

Why is it better to use a 25-base primer than a 10-base primer? Why do you have to have a lower annealing temperature for a 10-base primer?

Thank I appreciate it if you could help with these three questions. Ran out of questions to ask

Thank you again.

Explanation / Answer

Complementary sequence and annealing temperature controls the specific sequence to which PCR primer binds.

primer length and the GC content ( GC content means Guanine and Cytosine percentage or content in primer sequence.)

to get extra bands we have to change thermal cycler protocol and it would be decrease annealing temperature ( because higher annealing temperature lead to highly specific annealing but at lower temperature primer can bind to non specific sequence too and gives more bands.

If, after performing electrophoresis, there were no bands on the gel then we have to decrease annealing temperature of primer

why is it better to design your primers to complement an exon because econs are protein coding region of DNA and it is more reliable as well as efficient to amplification of target region.

it better to use a 25-base primer than a 10-base primer because shorter primer can bind to nonspecific sequence and gives wrong results,

to have a lower annealing temperature for a 10-base primer because efficient annealing to target sequence.

Related Questions

Navigate

• Browse (All)
• Browse Y
• Subjects

Integrity-first tutoring: explanations and feedback only — we do not complete graded work. Learn more.