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DNA Sequencing by Microchip Electrophoresis Using Mixtures of High-and Low-Molar Mass Poly(NN-dimethylacrylamide) Matrices

机译:使用高和低摩尔质量聚(NN-二甲基丙烯酰胺)基质的混合物通过芯片电泳进行DNA测序

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摘要

Previous studies have reported that mixed molar mass polymer matrices show enhanced DNA sequencing fragment separation compared to matrices formulated from a single average molar mass. Here, we describe a systematic study to investigate the effects of varying the amounts of two different average molar mass polymers on the DNA sequencing ability of poly(N,N-dimethylacrylamide) (pDMA) sequencing matrices in microfluidic chips. Two polydisperse samples of pDMA, with weight-average molar masses of 3.5 MDa and 770 kDa, were mixed at various fractional concentrations while maintaining the overall polymer concentration at 5% (w/v). We show that although the separation of short DNA fragments depends strongly on the overall solution concentration of the polymer, inclusion of the high-molar mass polymer is essential to achieve read lengths of interest (> 400 bases) for many sequencing applications. Our results also show that one of the blended matrices, comprised of 3% 3.5 MDa pDMA and 2% 770 kDa pDMA, yields similar sequencing read lengths (> 520 bases on average) to the high molar mass matrix alone, while also providing a five-fold reduction in zero-shear viscosity. These results indicate that the long read lengths achieved in a viscous, high molar mass polymer matrix are also possible to achieve in a tuned, blended matrix of high and low molar mass polymers with a much lower overall solution viscosity.
机译:先前的研究报道,与由单一平均摩尔质量配制的基质相比,混合摩尔质量的聚合物基质显示出增强的DNA测序片段分离。在这里,我们描述了一项系统研究,以研究改变两种不同平均摩尔质量聚合物的量对微流控芯片中聚(N,N-二甲基丙烯酰胺)(pDMA)测序基质的DNA测序能力的影响。将重均摩尔质量为3.5 MDa和770 kDa的两个pDMA多分散样品以不同的分数浓度混合,同时将总聚合物浓度保持在5%(w / v)。我们显示,尽管短DNA片段的分离在很大程度上取决于聚合物的整体溶液浓度,但对于许多测序应用而言,高分子量聚合物的包含对于实现感兴趣的读取长度(> 400个碱基)至关重要。我们的结果还表明,由3%的3.5 MDa pDMA和2%的770 kDa pDMA组成的混合基质之一,与单独的高摩尔质量基质产生的测序读长度(平均> 520个碱基)相似,同时还提供了五个零剪切粘度降低了两倍。这些结果表明,在高,低摩尔质量聚合物的调合共混基质中,在整体溶液粘度低得多的情况下,在粘性高摩尔质量聚合物基质中获得的长读取长度也是可能的。

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