Sensing and Actuation in Miniaturized Systems

PRESENTATION

DNA Transformation by Local Heat Shock

9533071 陳俊廷

Outline

Abstract Introduction Design And Fabrication Experimental Procedure Results Conclusions

Abstract

DNA transformation by local heat shock using a MEMS device has been successfully demonstrated using an on-chip micro heater and corresponding microfluidic system

The transformation experiments have been conducted and 200% improvement in transformation efficiency was showed as compared with control tests using conventional methodology

Introduction

Widely used in genetic engineering and molecular biology as an indispensable and key process in gene cloning, gene therapy and protein expression

Among all the DNA transformation methods, electroporation and heat shock are two commonly used procedures. Heat shock devices are easy to make and biocompatible.

Some researchers believe that the abrupt heat pulse makes cell membranes more fluid and permeable for the foreign DNA to transport into cells

1. Bacterial cells, which were treated by chemicals such as CaCl2 to make the cell membrane permeable

2. Plasmid DNA, a circular DNA that can replicate itself after DNA transformation process

Design and Fabrication

The competent cells and the recombinant plasmids are mixed

The mixture is then treated by a brief heat pulse at 42℃ for 90seconds for the transformation process

A fluidic cap using molded PDMS to construct micro chamber and channels and a heating substrate with microheaters constructed by resistive heaters with electrical insulation layer on the top

Experimental Procedure

A fine thermocouple (CHAL-0005), was good linearity between 0 to 65℃, TCR of the heater 0.0015/℃ and the coefficient of determination of the linear fit is 0.9999

E. coli stored was thawed on ice and mixed with plasmid DNA, which contains both Venus and ampicillin resistance gene

Compare local heat shock, conventional heat shock, and without heat shock

Results

The micro device achieves 2.62×106colonies/μg, which is about over 200% better than the conventional heat shock result at 1.13×106colonies/μg

No transformed cell can be observed without heat shock procedure

Conclusions

A MEMS local heat shock device has been successfully designed, fabricated and tested to demonstrate effective DNA transformation using competent E. coli cells and plasmid DNA with embedded Venus gene.

The improvement in efficiency is likely coming from the effective heating process in the micro scale . As such, this MEMS device has potentials to be part of lab-on-a-chip systems.