| Abstract Detail
Recent Topics Posters Baker, Stokes S. [1], Vidican, Cleo [2], Greib, Haittam [2], Jarocki, Christine [2], Stevens, Ryan [2], Roytek, Margaret A. [3]. An inexpensive epifluorescent attachment for digital cameras suitable for undergraduate laboratory experimentation with transgenic Arabidopsis expressing red-sifted GFP. An inexpensive epifluorescent attachment for digital cameras has been developed to photograph red-shifted Green Fluorescent Protein (GFP) expression in whole Arabidopsis plants. The apparatus was designed to support quantitative inquiry instructions. Formal assessments of undergraduates that used transgenic Arabidopsis plants that contained an environmentally regulated GUS reporter gene indicated that students increased their quantitative analytical skills after completing inquiry investigations (http://abstracts.aspb.org/pb2006/public/P23/P23029.html). However, the GUS reporter gene is not ideal for teaching laboratories because of the time-lag to detect gene expression and the qualitative nature of histochemical stains. Fluorescence in transgenic Arabidopsis expressing chimeric sGFP(S65T) genes (CaMV 35S promoter and Arabidopsis cor78 promoter) has been photographed with a Canon EOS 30D digital camera with an attached filter cube constructed with commercially available dichroic filters (magenta for excitation, yellow plus cyan for barrier, and 45o blue for beam splitting) that was illuminated with a 5 watt Luxeon V Star 440-460 nm blue light emitting diode. Tiff files were quantified with NIH ImageJ. Experiments with purified eGFP (BioVision) produced a linear camera response with protein concentration (r2 = 0.986) and exposure time (r2 = 0.981) when mean pixel intensity was below 35100. eGFP concentrations as low as 1.33 ng/μl produced a signal 3.2 fold above background. A 10 sec. exposure time produced the optimum signal to noise ratio. Chlorophyll fluorescence with the cyan filter removed produced curvilinear relationships for both concentration (r2 = 0.944) and exposure time (r2 = 0.984).  Fall 2007, a comparison study of the instructional effectiveness of the GFP expressing and GUS expressing plants will be conducted at the University of Detroit Mercy. Funded by National Science Foundation DUE grant 0442771.
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1 - University of Detroit Mercy, Biology Department, 4001 W. McNichols Rd., Detroit, MI, 48221, USA
2 - University of Detroit Mercy, Biology Department
3 - Oakland University, Department of Human Resource Development, School of Education and Human Services, Rochester, Michigan, 48309
Keywords:
Digital images
GFP
Epifluorescent
Inquiry
teaching
LED
light emitting diode
ImageJ
NIH Image
transgenic
Quantitative.
Presentation Type: Recent Topics Poster
Session: P
Location: Exhibit Hall (Northeast, Southwest & Southeast)/Hilton
Date: Sunday, July 8th, 2007
Time: 8:00 AM
Number: P79052
Abstract ID:2732 |
