Laser-induced heating for in situ DNA replication and detection in microchannels

Abstract

This study proposes a method for in situ local deoxyribonucleic acid (DNA) replication and detection in a long DNA strand through laser-induced heating and strong avidin-biotin binding. To achieve the target DNA replication, dielectrophoresis was generated to stretch and immobilise DNA strands on both ends of the electrode. Subsequently, local DNA sequences were replicated using thermal cycles generated by laser-induced heating. Replicated double-stranded DNA products were captured in situ on a solid surface and detected using the fluorescence intensity of quantum dots (Qdots). The results revealed that after six laser-induced thermal cycles, the replicated local DNA sequence could be detected by analysing the difference between Qdot fluorescent intensity before and after replication. The proposed method is expected to improve the efficiency of biosample gene sequence analysis.

Fig. 1

Schematic describing the process of in situ replication and detection of the DNA strand

Fig. 2

Schematic of the experimental set‐up of the microchannels (a) Schematic diagram of the Al electrodes, (b) The experimental system set‐up

Fig. 3

Relation between laser power and temperature using fluorescence‐based temperature measurement

Fig. 4

Fluorescent images (photos were artificially brightened) of Qdots on the microchannel surfaces (a) Without BSA, (b) Coating with BSA

Fig. 5

Fluorescent images of the stretch and immobilised DNA on the electrode (a) After electric field applied, (b) After fluorescent dye injection

Fig. 6

Fluorescent images of the stretch and immobilised DNA on the electrode (a) Before laser irradiation, (b) After laser irradiation

Fig. 7

Fluorescent images (photos were artificially brightened) of the Qdots on the surface after thermal cycles through laser‐induced heating (a) The stretch DNA, (b) Six thermal cycles, (c) Nine thermal cycles, (d) Twelve thermal cycles, (e) Fifteen thermal cycles, (f) Eighteen thermal cycles

Fig. 8

Calculated result of fluorescent intensity at various thermal cycles through laser‐induced heating

Fig. 9

Results of DNA replication (30 cycles) by PCR machine Lane 1, standard sample; lane 2, recovery sample after laser heating 6 cycles; lane 3, recovery sample after laser heating 9 cycles; lane 4, recovery sample after laser heating 12 cycles; lane 5, recovery sample after laser heating 15 cycles; lane 6, recovery sample after laser heating 18 cycles; lane M, marker