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BISC110/F12: Series 3 Experiment 9 Hill Reaction: Difference between revisions
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BISC110/F12: Series 3 Experiment 9 Hill Reaction (view source)
Revision as of 12:33, 7 May 2012
, 7 May 2012→Procedure
Tucker Crum (talk | contribs) (New page: {{Template:BISC110/F12}} <div style="padding: 10px; width: 725px; border: 5px solid #1919FF;"> =='''Series 3- Lab 9 Photosynthesis -Photosynthetic Pigments and The Hill Reaction'''== In to...) |
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'''Materials:''' Each pair will have a ringstand with a test tube holder, and a 150W quartz-halogen projection lamp. You will have to find the best arrangement to measure the reduction of DCPIP accurately and reproducibly. A light meter and timer will be provided. Set up 5 ‘reaction tubes’ to begin with, adding 5 mL of '''reaction mixture''', but do not add the thylakoid membranes, which must be kept on ice. Add these thylakoid membranes just before each reaction mixture is tested. [Reaction mix: 50 mM sodium phosphate, pH 6.8 buffer, 100 mM sorbitol, 5 mM MgCl2, 0.05 mM DCPIP]<br><br> | '''Materials:''' Each pair will have a ringstand with a test tube holder, and a 150W quartz-halogen projection lamp. You will have to find the best arrangement to measure the reduction of DCPIP accurately and reproducibly. A light meter and timer will be provided. Set up 5 ‘reaction tubes’ to begin with, adding 5 mL of '''reaction mixture''', but do not add the thylakoid membranes, which must be kept on ice. Add these thylakoid membranes just before each reaction mixture is tested. [Reaction mix: 50 mM sodium phosphate, pH 6.8 buffer, 100 mM sorbitol, 5 mM MgCl2, 0.05 mM DCPIP]<br><br> | ||
'''Blanking the Spectrophotometer '''([[Appendix_B/ | '''Blanking the Spectrophotometer '''([[Appendix_B/F12 | Appendix B]]): Use a tube containing 5 mL of '''resuspension solution''' and 50µL of your '''thylakoid suspension''' to blank the Spec20 at 580 nm (filter level to the left). With Parafilm® over the top of the tube, invert once to mix, then wipe the tube with a Kimwipe® before blanking the instrument. Do not vortex. You may need to prepare a fresh blank periodically. <br><br> | ||
'''Varying the Reaction Rate:''' The Hill reaction occurs at different rates depending on the light intensity supplied to the thylakoids. You will measure the reaction rates by taking absorbance readings over a 90 s period in 15 s intervals. We will use the drop in absorbance over time as a measure of the rate of the Hill reaction. <br><br> | '''Varying the Reaction Rate:''' The Hill reaction occurs at different rates depending on the light intensity supplied to the thylakoids. You will measure the reaction rates by taking absorbance readings over a 90 s period in 15 s intervals. We will use the drop in absorbance over time as a measure of the rate of the Hill reaction. <br><br> | ||
Depending on the number, distribution (clumped or not) and condition of thylakoids in your reaction tube, you may obtain varying reaction rates with a given light intensity. We measure the incident light using a light meter in units of µmol photons m-2 s-1. <br><br> | Depending on the number, distribution (clumped or not) and condition of thylakoids in your reaction tube, you may obtain varying reaction rates with a given light intensity. We measure the incident light using a light meter in units of µmol photons m-2 s-1. <br><br> | ||
