Shear-induced degradation of linear polyacrylamide solutions during pre-electrophoretic loading.
Electrophoretic channels are filled with a polymer matrix prior to their use in DNA separations. This process, called gel-loading, can be accomplished manually, using syringes, or can be automated through the use of small pumps or vacuum. The injection rate is constrained by the desire to minimize shear-induced degradation of the polymer molecules. Currently, the community lacks quantitative data with which to gauge the range of flow rates that prevent polymer degradation. In this study, measurements of the zero shear rate viscosity of linear polyacrylamide (LPA) solutions are used to determine the LPA molecular weight before and after gel-loading. The results indicate molecular degradation in polymer solutions even when injected at minimal flow rates of 1 microL/min. To correlate these rheological observations of shear-induced degradation with subsequent electrophoretic performance, the degraded solutions were used as sieving matrixes for DNA sequencing analysis. The decreases in electrophoretic resolution and increases in peak widths between sheared and nonsheared LPA solutions are related to the degradation in molecular weight experienced by the polymer solutions.
Vazquez
M
M
Whitehead Institute for Biomedical Research, Cambridge, Massachusetts 02142, USA.
Schmalzing
D
D
Matsudaira
P
P
Ehrlich
D
D
McKinley
G
G
D002138:Calibration; D004247:DNA / Q000302:isolation & purification; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods*; D011134:Polysaccharides / Q000737:chemistry*; D012996:Solutions
D011134:Polysaccharides; D012996:Solutions; D004247:DNA