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在一次性使用的微加工硅芯片上对基因组DNA中的2千碱基蓝藻基因和人抗α1-糜蛋白酶基因进行聚合酶链反应。

Polymerase chain reaction of 2-kb cyanobacterial gene and human anti-alpha1-chymotrypsin gene from genomic DNA on the In-Check single-use microfabricated silicon chip.

作者信息

Consolandi Clarissa, Severgnini Marco, Frosini Andrea, Caramenti Giancarlo, De Fazio Marco, Ferrara Francesco, Zocco Anna, Fischetti Alessandra, Palmieri Michele, De Bellis Gianluca

机构信息

Institute of Biomedical Technologies, National Research Council, ITB-CNR, Via Fratelli Cervi 93, I-20090 Segrate, MI, Italy.

出版信息

Anal Biochem. 2006 Jun 15;353(2):191-7. doi: 10.1016/j.ab.2006.03.019. Epub 2006 Mar 31.

Abstract

The microfabricated chip is a promising format for automating and miniaturizing the multiple steps of genotyping. We tested an innovative silicon biochip (In-Check Lab-on-Chip; STMicroelectronics, Agrate Brianza, Italy) designed for polymerase chain reaction (PCR) analysis of complex biological samples. The chip is mounted on a 1x3-in(2). plastic slide that provides the necessary mechanical, thermal, electrical, and fluidic connections. A temperature control system drives the chip to the desired temperatures, and a graphical user interface allows experimenters to define cycling conditions and monitor reactions in real time. During thermal cycling, we recorded a cooling rate of 3.2 degrees C/s and a heating rate of 11 degrees C/s. The temperature maintained at each thermal plateau was within 0.13 degrees C of the programmed temperature at three sensors. From 0.5 ng/microl genomic DNA, the In-Check device successfully amplified the 2060-bp cyanobacterial 16S rRNA gene and the 330-bp human anti-alpha(1)-chymotrypsin gene. The shortest PCR protocol that produced an amplicon by capillary electrophoresis comprised 30 cycles and was 22.5 min long. These thermal cycling characteristics suggest that the In-Check device will permit future development of a genotyping lab-on-a-chip device, yielding results in a short time from a limited amount of biological starting material.

摘要

微制造芯片是一种很有前景的形式,可实现基因分型多个步骤的自动化和小型化。我们测试了一种创新的硅生物芯片(In-Check芯片实验室;意法半导体公司,意大利阿格拉特布里安扎),该芯片专为对复杂生物样本进行聚合酶链反应(PCR)分析而设计。芯片安装在一块1×3英寸(2)的塑料载玻片上,该载玻片提供必要的机械、热、电和流体连接。温度控制系统将芯片驱动至所需温度,图形用户界面允许实验人员定义循环条件并实时监测反应。在热循环过程中,我们记录到冷却速率为3.2℃/秒,加热速率为11℃/秒。在三个传感器处,每个热平台维持的温度与设定温度的偏差在0.13℃以内。从0.5 ng/μl基因组DNA开始,In-Check设备成功扩增了2060 bp的蓝藻16S rRNA基因和330 bp的人抗α(1)-糜蛋白酶基因。通过毛细管电泳产生扩增子的最短PCR方案包括30个循环,时长为22.5分钟。这些热循环特性表明,In-Check设备将有助于未来基因分型芯片实验室设备的开发,能在短时间内从有限量的生物起始材料中得出结果。

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