连续流动式PCR微流控装置的研究

A Study on a Continuous-flow PCR Microfluidics

介绍了基于薄膜加热器的新型连续流动式PCR微流控装置的设计与制作;讨论了退火温度、PCR反应试剂(引物、Mg2+、dNTPs以及Taq DNA聚合酶)浓度以及PCR溶液的流动速度等对连续流动式PCR反应的影响;结果发现反应试剂影响连续流动式微流控PCR扩增的行为不同于它们影响传统PCR的行为,在较宽的浓度范围内都不会引起非特异性扩增。除此之外,在15 min内能成功对249 bp的人类β-肌动蛋白基因进行扩增,扩增速度比传统PCR快;通过低热容量的薄膜加热器来维持三个温度区带的恒温,完成33个循环的连续流动式PCR扩增能量消耗小于0.0088 kW.h,比传统PCR仪低得多,新研制的PCR微流控装置有可能成为便携式装置。

A new continuous-flow PCR microfluidics was successfully designed and constructed, in which several factors that might affect the continuous-flow PCR amplification had been investigated in detail, including annealing temperature, concentrations of some PCR reagents （forward/reverse primers, Mg^2＋ , dNTPs and DNA polymerase enzyme） and flow rates of PCR solution in the capillary microchannel. It has been found that the ways these reagents affect the continuous-flow PCR are different from those that affect the conventional PCR, and that these reagents within a wider concentration range wont result in the non-specific PCR amplification. In addition, the 249 bp β-actin gene fragments could be successfully amplified on the present PCR microfluidlcs within 15 mins, which was much faster than the reaction time with the conventional PCR machine. Using the flexible thin film heaters with low thermal mass to maintain the three thermostable zones, the energy consumption of a 33 continuous-flow PCR cycles is less than 0. 0088 kW · h, which may make the present continuous-flow PCR microfluidics portable.