Molecular Vision: Ryan, Mol Vis 2016; 22:1169-1175. Table 1

Table 1. PCR primers used in the diagnostic test.

Mutation	Primer (5’-3’)	Product size (Position)	Melt temperature average (range)
3460A ARMS	F: TACTACAACCCTTCGCTGcCA*	120 bp (3440–3559)	82.58 °C (82.32 - 83.14)
	R: GTAGAAGAGCGATGGTGAGAGCTAAG
11778A ARMS	F: ACGAACGCACTCACAGTgA	143 bp (11,760-11902)	80.32 °C (80.08 - 80.7)
	R: CACAGAGAGTTCTCCCAGTAGGTTAA
14484C ARMS	F: GTAGTATATCCAAAGACAACgAC*	161 bp (14,462-14622)	76.31 °C (76.01 - 76.77)
	R: GGGTTTTCTTCTAAGCCTTCTCC
Insulin control	F: GACTGAATTCCGGCACGTCCTGGCAGTG	221 bp (1458–1658)	90.24 °C (90.04 - 90.68)
	R: GACTCTCGAGGGAGGCGGCGGGTGTG

The bases highlighted in bold result in allele specific amplification of the 3460A, 11778A and 14484C mutated sequences respectively. Lower case bases highlight mismatches to increase the specificity of the allele specific amplification. The PCR product sizes, positions in Homo sapiens mitochondrion, complete genome (NCBI Reference Sequence: NC_012920.1) or genomic insulin sequence (E00022.1) and average melting temperatures are highlighted. The melting temperatures are based on at least 9 separate experiments. The 2 primers marked with * have previously been published by Bi et al. 2010 [30].