Application of Quantum Dot Labeling to DNA Microarray
Date Issued
2008
Date
2008
Author(s)
Jian, Zhen-Zhi
Abstract
DNA Microarray technology is extremely powerful for detecting gene expression in large scale. The accuracy and precision of microarray depends on the fluorescent signal of fluorescent labeled cDNA or cRNA probe. Netherless, conventional organic fluorophores are easily photobleaching. The most used dyes, cyanine 3 and cyanine 5, are affected by ozone, especially cyanine 5. An alternative fluorophore for labeling are required for DNA microarray analysis.he aim of this thesis was to establish and optimize a quantum dot labeling system for DNA microarray. In this thesis, I present a microarray system based on quantum dot labeling method, in which cDNA were labeled with biotin and hybridized with 60 mer oligo probe on the glass slide. After incubation with quantum dot streptavidin conjugated, the fluorescent signal intensity of the quantum dot streptavidin conjugated bound to the biotin labeled cDNA detected by a laser scanner. found that quantum dots are more photostable than Cy5-dUTP at ambient air. The bluing effect of streptavidin conjugated Qdot®655 was not obvious at ambient air within one hour. Four incubation conditions, two kinds of biotin labeling methods and two kinds of coated slides were testified to optimize the system. Besides, I used a lung cancer cell line-CL1-5 total RNA and a housekeeping gene, Glyceraldehyde-3- phosphate dehydrogenase, 60mer probe as RNA and probe in model microarrays. The detection dynamic range are about 2 orders of magnitude.
Subjects
Quantum dots
DNA microarray
Hybridization
SDGs
Type
thesis
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