Post on 31-Dec-2015
Redefining DNA Microarrays
Tiffany McAninchAdvised by:Dr. Frederick Haselton, PhDMark McQuain, 2BPhD
Purpose
If humanity was able to understand genetics more completely and able to determine what the causes were for many disorders, there would be a huge sweep for this information. DNA microarrays are the most recent tool to put this information to use in a large scale way. This project has been to design a system similar to microarrays, but one in which the information can be obtained faster, more accurately, and more easily.
Northern Blotting
Microarray Procedure
Target genes prepared and hyb. onto slides
Culture cellsIsolate mRNAPrepare labeled cDNA2 cDNAs competitively hybridizedAnalyzed by fluorescent signature
Microarray Schematic
mRNA from Sample 1 mRNA from Sample 2
Reverse transcribe each sampleusing a different fluorescent
nucleotide (CY3 or CY5)
Mix equal amounts of eachcomplex probe together and
hybridize overnight
Scan and quantitate gene expression levels
(Green Fluorescence) (Red Fluorescence)
Streptavidin Beads
Flow Cytometer
PMT
PMT
PMT
Dichroic Filters
BandpassFilters
Laser
Flow Manifold
SSC
FSC
Cells
Raman Spectroscopy
Trp-Phe
0.00E+00
2.00E+03
4.00E+03
6.00E+03
8.00E+03
1.00E+04
0 500 1000 1500 2000
Rel cm
Ram
an I
nte
nsi
ty
Phe-Gly-Phe-Gly
0.00E+00
5.00E+02
1.00E+03
1.50E+03
2.00E+03
2.50E+03
0 500 1000 1500 2000
Rel cm
Ram
an I
nte
nsi
ty
Trp-Trp
0.00E+00
5.00E+02
1.00E+03
1.50E+03
2.00E+03
0 500 1000 1500 2000
Rel cm
Ram
an I
nte
nsi
ty
Comparison
Microbeads Easier to store Do chemically Hold patent! Less Reagents Speed Flow Cytometer 2 million signatures
Microarrays Don’t need Raman
tag No flow cyt. for
Raman 50,000 signatures
Conclusion
The design has been formulated, which was the purpose of this project. There has also been much research that will aid in future use of narrowing the design further. Having found the beads with suitable surface chemistries and measured the Raman of the tags, the project is ready to be further tested.
Thanks!!
Professor HaseltonMark McQuainProfessor Mahadevan-JansenChad LieberTodd Monroe