DNA barcoding via counterstaining with AT/GC sensitive ligands in injection-molded all-polymer nanochannel devices

Abstract

Nanochannel technology, coupled with a suitable DNA labeling chemistry, is a powerful approach for performing high-throughput single-molecule mapping of genomes. Yet so far nanochannel technology has remained inaccessible to the broader research community due to high fabrication cost and/or requirement of specialized facilities/skill-sets. In this article we show that nanochannel-based mapping can be performed in all polymer chips fabricated via injection molding: a fabrication process so inexpensive that the devices can be considered disposable. Fluorescent intensity variations can be obtained from molecules extended in the polymer nanochannels via chemical counterstaining against YOYO-1. In particular, we demonstrate that the counterstaining induced fluorescent intensity variations to a large degree appear to be proportional to the theoretically computed sequence-maps of both local AT and GC variation along DNA sequences.

Surface roughness reduction using spray-coated hydrogen silsesquioxane reflow

“…Roughness of advanced polymer replication tools is costly and challenging to control, especially in case of complex geometries such as tools for freeform optical components, for microfluidic and medical diagnostics devices [1][2][3] or as substrates for nanopatterned surfaces. Such applications may often require special fabrication of molding tools.…”

Polymeric-Based In Vitro Diagnostic Devices

“…Injection molding has been used to produce LOC systems for on-chip liquid chromatography and DNA bar coding systems [43,80]. However, this process can be challenging because of the large aspect ratios required, low surface roughness, high flow pattern resolution, and stresses experienced by the replica.…”

“…Replication of nanostructures into polymer surfaces has been successfully achieved and had received a good deal of attention ( Fig. 2.3) [80,81]. The main limitation of this process lies in the grain size of the master, which limits its ability to successfully replicate smaller structures.…”

“…Among them, SU-8 photoresist has also been used as a master material through standard photolithography for embossing polymeric materials under conditions similar to the ones used with metal templates, and an aluminum coating can be applied to facilitate substrate demolding from the template [87]. To fabricate high aspect ratio structures, an [80] anodic aluminum oxidation nanostructured membrane and cylindrical pillars can be used [88,89].…”

DNA translocation through short nanofluidic channels under asymmetric pulsed electric field

“…Examples of such biological phenomena include passage of mRNA through nuclear pore complexes, injection of DNA from a virus head into a host cell, and protein translocations across biological membranes through channels. 1 The study of DNA as a model polymer in confined environments, such as nanopores, 2-4 nanochannels, [5][6][7][8][9] and nanoslits, [10][11][12][13][14][15] has led to significant advancements in our understanding of DNA dynamics 11,[16][17][18] and its application to DNA sorting, 10,12,[19][20][21] mapping, [22][23][24][25] and sequencing. 2,26 DNA translocation through a nanochannel can be viewed as a three stage process-first, one end of the polymer chain reaches the nanochannel entrance, then the chain enters the nanochannel, and finally the chain exits the nanochannel.…”