Question

21) You have this small piece of sequence data from a portion of your favorite gene that was isolated fronm a ladybug cDNA library. The spaces are to help with counting and the part not show in the midle () is 263 bp. 5' tca tta tgg tgt ttg agt aca tga aac acg gag atc tca ac.. agg tgo gaa tgg cag acg gac age aga gtt 3' a) Design forward and reverseprimersfor PCR that will isolate EXACTLY the sequence that is in bold typeface. Guidelines Make your primers 18 bp long Write your primers in 5' to 3' direction Forward 5' Reverse 5 3' 3' You get your primers, and you test them in a PCR reaction on your cDNA library. b) What size band do you expect when you run out the PCR products out on an agarose gel? bp

Answer 1

a)

While designing primers, the sequence of the forward primer is same as that of the region to be amplified and the sequence of the reverse primer is complimentary to the strand when read from 3' - 5' direction. Thus, the sequence of forward and reverse primer would be :-

Forward : 5'-ttgagtacatgaaacacg- 3'

Reverse : 5'- gctgtccgtctgccattc -3'

b)

The band size of the PCR products would be the length of the gene/sequence to be amplified. In this case, it would be 323 bp.