Rapid screening mitochondrial DNA mutation by using denaturing high-performance liquid chromatography

Table 1 Primer sequence, PCR Conditions and DHPLC oven temperature used in this study

Fragments	Size (bp)	Primer Sequence	Annealing temp. (°C)	DHPLC temp. (°C)	DHPLC start gradient
A3243G	494	F: CCTCCCTGTAGGAAAGGACA	59	58	55% Ba
		R: GCCTAGGTTGAGGTTGACCA
D-loop F1	505	F: GCTGGAAGATCTTTAACTCCACCATTAGCACC	65	58	58% B
		R: CTACGCGTCGACGCGAGGAGAGTAGCACTCTTG
D-loop F2	515	F: GCTGGAAGATCTAAATCAATATCCCGCACAAG	65	58	58% B
		R: CTACGCGTCGACTTAAGTGCTGTGGCCAGAAG
D-loop F3	494	F: GCTGGAAGATCTCACCCTATTAACCACTCACG	58	57	58% B
		R: CTACGCGTCGACTGAGATTAGTAGTATGGGAG
D-loop F4	585	F: GCTGGAAGATCTACAAAGAACCCTAACACCAGC	65	58	59% B
		R: CTACGCGTCGACACTTGGGTTAATCGTGTGACC
16s-F2	292	F: ATATAGTACCGCAAGGGAAAGA	53	56	49% B
		R: GGGTTCTGTGGGCAAAT
16s-G6	268	F: AATAGGGACCTGTATGAATGG	53	60	51% B
		R: TAGTTCGCTTTGACTGGTG

^aEluent buffer B was 0.1 mmol·L^-1 TAEE, 25% acetonitrile, pH7.0.

Table 2 The detected frequency of variance of mtDNA by using DHPLC

Fragments Samples	A3243G			D-loop
	MELAS	Other Encephala	Normal blood	Tumor	Cell line
Positive/total fragments	10/12	0/72	0/30	26/184	38/52
Percent (%)	83.3	0	0	14.1	73.1