User:Elaine Marie Robbins/Notebook/CHEM-496/2011/09/27: Difference between revisions
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# Combine 10 mL of 50 mM acetate buffer (pH 5.46) with 0.974 mL of 0.0015 mM BSA and 0.581 mL of 0.25 mM HAuCl<sub>4</sub> in a capped test tube at room temperature. | # Combine 10 mL of 50 mM acetate buffer (pH 5.46) with 0.974 mL of 0.0015 mM BSA and 0.581 mL of 0.25 mM HAuCl<sub>4</sub> in a capped test tube at room temperature. | ||
# Take UV vis spectra (200 nm to 800 nm) of a sample of reaction mixture at 70°C every 30 min. | # Take UV-vis spectra (200 nm to 800 nm) of a sample of reaction mixture at 70°C every 30 min. | ||
# Place reaction mixture in an oven at 80°C for 2.5 hr under thermostatic conditions. | # Place reaction mixture in an oven at 80°C for 2.5 hr under thermostatic conditions. | ||
# After 2.5 hr, remove from oven and cool to room temperature; leave sitting overnight. | # After 2.5 hr, remove from oven and cool to room temperature; leave sitting overnight. | ||
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# [http://openwetware.org/wiki/AU_Biomaterials_Design_Lab:Protocols/Autoclave| Autoclave]. | # [http://openwetware.org/wiki/AU_Biomaterials_Design_Lab:Protocols/Autoclave| Autoclave]. | ||
# Allow flask to cool. | # Allow flask to cool. | ||
# Add 35 μL of | # Add 35 μL of 1000× ampicillin. | ||
# Inoculate with the bacteria you are culturing by scraping frozen bacteria/glycerol mixture and adding to flask. | # Inoculate with the bacteria you are culturing by scraping frozen bacteria/glycerol mixture and adding to flask. | ||
# Place the flask in a shaker/incubator and culture overnight at 37°C and 200 rpm. | # Place the flask in a shaker/incubator and culture overnight at 37°C and 200 rpm. | ||
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# Make LB/agar plates with ampicillin. | # Make LB/agar plates with ampicillin. | ||
# Place sterile tube and DNA in ice bucket for 15 min. | # Place sterile tube and DNA in ice bucket for 15 min. | ||
# Add 5 μL of DNA and | # Add 5 μL of DNA and 40 μL of cells to the bottom of the tube. | ||
# Incubate on ice for 30 min. | # Incubate on ice for 30 min. | ||
# Heat shock DNA/cells at 42°C for 30 s. | # Heat shock DNA/cells at 42°C for 30 s. |
Revision as of 10:03, 19 October 2011
AuNP III, Expression, DNA Transformation | <html><img src="/images/9/94/Report.png" border="0" /></html> Main project page <html><img src="/images/c/c3/Resultset_previous.png" border="0" /></html>Previous entry<html> </html>Next entry<html><img src="/images/5/5c/Resultset_next.png" border="0" /></html> |
ObjectiveAuNP III To synthesize gold nanoparticles (AuNP) using the procedure developed here:
http://pubs.acs.org/doi/abs/10.1021/jp110296y
To begin culturing bacteria to express inteins.
To transform DNA coding for mutant GFP (mutated on 09/20/11) into Novablue cells. DescriptionAuNP III
Starter Culture Media, Protein Expression
DataAuNP III The absorbance values for the AuNP solution were subtracted from the absorbance values for a blank acetate buffer to create a set of corrected absorbance values. The corrected absorbance values at 550 nm were graphed against time. The graph shows a positive linear relationship, indicating that AuNPs were formed in solution. In addition, a second graph showing corrected absorbance against wavelength was created, to show the absorbance levels across all of the wavelengths measured by the UV-Vis. A magnified version of this graph shows that absorbance values at around 550 nm increased over time, indicating the creation of AuNPs.
Plate did not appear to be successful; no cells grew. NotesThe AuNP solution turned from clear to purple while in the oven, indicating that AuNPs were actually formed.
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