Separation of DNA fragments by capillary electrophoresis using replaceable linear polyacrylamide matrices

Is Higher Concentration of Linear Polyacrylamide Not Suitable for the Capillary Electrophoretic Separation of Large DNA?

“…l-4,9,10 Linear polyacrylamide has been used as a separation matrix for DNA sequencing especially for long reading by capillary electrophoresis. 5-g In our common knowledge, higher concentration of polyacrylamide is suitable for separating small DNA fragments and lower concentration of polyacrylamide is indispensable for the separation of large DNA fragments.11-13 Low concentration of polyacrylamide gel (3% T and 0.5% C) is applied to the separation of up to 12 kbp DNA fragments1,2,4 and 1.5% linear polyacrylamide gave a fine resolution of DNA fragments up to 23 kbp3, whereas shorter DNA fragments up to 1 kbp should be separated by using 6% linear polyacrylamide.l, 3 Large DNA fragments up to 48.5 kbp could only be separated by using very diluted linear polyacrylamide (0.6%).9 Use of lower concentration of linear polyacrylamide (2%) allowed DNA sequencing of more than 1000 bases per rung, whereas DNA sequencing of less than 500 bases was achieved using 6% linear polyacrylamide as a separation matrix. 5,6 More recently, some cellulose derivatives were proven to be an efficient separation matrix for DNA fragments.l3-19 In this instance, use of ultradilute polymer solution (0.00125%) was necessary for the separation of DNA up to 23 kbp17-19, whereas 0.5% cellulose solution is suitable for the separation of DNA fragments up to 1 kbp.14-16 A pulsed field technique20 coupled with use of ultradilute polymer solution gave an excellent separation of multikilobase and mega base DNA fragment2l-26, which could not be separated without a pulsed filed technique.…”

Intrinsic isotachophoretic preconcentration in capillary gel electrophoresis of DNA restriction fragments

“…Linear polyacrylamide gel buffers were prepared by polymerisation of 8% acrylamide dissolved in 0.1 M Tris buffer containing 0.5% TEMED and 0.08% ammonium persulphate for 48 h at 4°C [12]. The Tris buffer was adjusted to pH 8.3 with hydrochloric acid, boric acid or benzoic acid.…”

Polymer solution-filled column for the analysis of antisense phosphorothioates by capillary electrophoresis

“…Although a linear polyacrylamide solution shows little viscosity at concentrations below 6% [25], a concentration above 10% is usually required for separating ODNs with a base length of less than 25. At this range of concentrations, the linear polyacrylamide solution becomes fairly viscous and difficult to be replaced or regenerated.…”