Nucleic acid stain-dependent single strand conformation polymorphisms

Figures & data

Figure 1.  Four single strand conformational polymorphism (SSCP) staining methods in a single polyacrylamide gel.

SSCP of H09g marker for two Tribolium castaneum strains (M2, mas p au; T2, Tiw-1) show duplicate applications in each of four treatments (EB, ethidium bromide; SYB, SYBR Gold). λ DNA (50 ng) loaded into lanes marked “L” served as fluorescent controls. This 10% polyacrylamide gel [99:1 methylene-bis-acrylamide in 1× Tris-borate EDTA (TBE)] was run in a vertical position at 10°C (5°C water jacket) with 0.5× TBE running buffer for 60 min at 180 constant volts. Pre- and post-stained samples were run on the same gel and physically separated for postelectrophoretic treatment.

Figure 2.  Pre-stain versus post-stain single strand conformational polymorphism (SSCP).

Negative fluorescent images are shown for two strains of Tribolium castaneum (M, mas p au; T, Tiw-1), two staining methods (SYBR Gold pre-stain, ethidium bromide post-stain), and nine molecular markers. Markers are grouped as: (A) stain-dependent polymorphisms (H09g, H115, M2s); (B) stain-independent polymorphisms (A011, B16s, F2s); and (C) nonpolymorphic (Tsh, J1s, and CHSA). Polyacrylamide gel electrophorsis conditions were same as Figure 1 and run for 90 min. Pre- and post-stained samples were run on the same gel and physically separated for postelectrophoretic treatment. STS, sequence-tagged site.

Table 1.  STS Polymorphisms

STS	Size (bp)	SNP (No.)	Indel (bp)
H09g	401	3
H115	138	1
M2s	317–320	2	3
A011	251	2
B16s	157–169	3	10,2
F2s	309–310	3	1
Tsh	236	0
J1S	142	0
CHSA	467	0

Shown are the size, number of single nucleotide polymorphisms (SNPs), and size of insertions/deletions (Indel) underlying SSCPs detected between the mas p au and Tiw-1 strains of Tribolium castaneum. STS H115 was designed to isolate a single SNP of STS H09g. STS, sequence-tagged site; SSCPs, single strand conformational polymorphism.