Branched polymeric labels used as drag-tags in free-solution electrophoresis of ssDNA

Abstract

In the framework of the classical blob theory of end-labeled free-solution electrophoresis of ssDNA, and based on recent experimental data with linear and branched polymeric labels (or drag-tags), the present study puts forward design principles for the optimal type of branching that would give, for a given total number of monomers, the highest effective frictional drag for ssDNA sequencing purposes. The hydrodynamic radii of the linear and branched labels are calculated using standard models like the freely jointed chain model and the Kratky-Porod worm-like chain model. Based on comparisons of the theory with the experimental data, we propose that the design of new branched labels should use either side chains whose length is comparable to the distance between the branching points or two long branches located near the ends of the molecule's backbone.

Theory of end‐labeled free‐solution electrophoresis: Is the end effect important?

“…For the situation when the DNA and the drag‐tag have different mechanical properties, we have only considered the extreme case when one of the parts is completely flexible and the other completely rigid. In previous works neglecting the end effect , the case of semiflexible parts with different persistence lengths was treated by dividing both parts into “blobs” of equal hydrodynamic radii, treating each blob as the new “effective monomer” and thus reducing the problem to that of a flexible chain with a constant monomer size. This reduction should still apply in principle even when the end effect is taken into account, but only if it is possible to subdivide both parts into blobs in such a way that both the number of persistence lengths per blob is large and the number of blobs is large as well.…”

“…For the latter, theories and computer simulations of electrophoresis of composite objects are necessary. Besides generic theories not making specific assumptions about the drag-tag [3] and the simplest case of a flexible diblock copolymer in a weak field [15][16][17][18], the problems considered theoretically so far include globular [15,19] and branched [18,20] drag-tags, separation and stretching of a drag-tag-DNA composite in a strong field [21][22][23][24], and dragtags attached to both ends of the DNA [25]. Besides ELFSE, such studies can also be useful for understanding other situations of practical interest involving objects consisting of two or more parts with different electrophoretic properties, for instance, different variants of affinity electrophoresis [26][27][28][29][30].…”

Quantitative SNP genotyping by affinity capillary electrophoresis using PEG‐oligodeoxyribonucleotide block copolymers with electroosmotic flow

“…In addition, Barron and coworkers reported that the electrophoretic mobility of the block-type conjugates between ssDNA and branched poly(N-methoxyethyl glycine) was dependent solely on the peptide's molecular weight, which ranged from 4K to 9K, regardless of its branch number [54]. The theoretical investigations on the phenomena were also conducted by Slater et al [55].…”

Molecular deformation and free‐solution electrophoresis of DNA‐uncharged polymer conjugates at high field strengths: Theoretical predictions. Part 1: Hydrodynamic segregation

“…ELFSE has been used to sequence up to about 100 base long ssDNA molecules in about 18 min in a 34 cm long capillary, using the globular protein streptavidin as a drag-tag [1]. Current research is directed at finding larger drag-tags, possibly with nonlinear architectures [12,13] to extend the read length of ELFSE [4].…”