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. 2013 May 28;8(5):e65141. doi: 10.1371/journal.pone.0065141

Figure 4. Reverse transcriptase-PCR for mutant and wild type KEAP1 .

Figure 4

(A) Primer positions for reverse transcriptase-PCR in KEAP1 exons. The asterisk indicates the position of c.879_880delinsA mutation in exon 3. The primer, KEAP1-mut-rvs, contains "T" nucleotide at the 3' end which is the complement of mutant "A" nucleotide to amplify mutant allele. The expected sizes of PCR products are as follows: 233 bp for KEAP1-N-fwd and KEAP1-N-rvs; 201 bp for KEAP1-C-fwd and KEAP1-C-rvs; 282 bp for KEAP1-mut-fwd and KEAP1-mut-rvs. (B) Results of the reverse transcriptase-PCR for the mutant and wild type KEAP1. cDNAs from control normal thyroid, the normal and goitre region of thyroid tissues obtained from the proband were amplified by 3 primer sets indicated in (A) to mutant KEAP1 (upper panel), 5' and 3' normal portions adjacent mutation site (second and third panels, respectively) of KEAP1. GAPDH was amplified as a control for the amount of cDNA in each sample. The expected size of PCR product of GAPDH is 452 bp. In the experiment of mutant KEAP1 amplification, the plasmids containing mutant or wild type KEAP1 (pCMV-myc-KEAP1mut or pCMV-myc-KEAP1wt, respectively) were utilized as control templates.