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Methods and Materials |
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| Two Chrysanthemum plants were purchased from Wal-Mart and remained in the same pots they were purchased with throughout the length of the experiment. The two plants were placed on a window sill in direct sunlight for 5 days. For this period, one of the plants, designated for desiccation, was not watered at any point. The other plant was watered with approximately 20mL of tap water every day at 6 PM. At the end of the five day period, leaves and flowers were collected from both plants. The leaves and flowers were collected from all sides of the plants. The samples were stored in plastic bags in a 4oC refridgerator until they were used. | |
| Western Blot |
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| The samples were removed from the fridge and protein was
extracted for the
Western Blot procedure, which will identify
and isolate RBCL protein.
Protein isolation was performed on all four of the plant samples and
DC protein assay was performed to determine the protein concentration for
the Western Blot. The protein was then kept frozen in the -70oC
freezer until it was used for the Western Blot. Once the Western Blot was performed,
antibody detection via a light reaction was used to identify RBCL. Note: A 1.5 mg/mL standard was used in place of the 1.25 mg/mL standard for the protein isolation. Note: The alternative antibody detection procedure was used. No milk was used in the preparation of blocking buffer, and the antibody dilutions were 1:20,000 for both the primary chicken antibody and the secondary goat anti-chicken antibody. |
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| DNA Isolation and Analysis |
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| DNA was
extracted from all four samples and concentrations were determined
using the Nanodrop.
Real-time PCR amplified the RBCL gene, which was then analyzed using
agarose gel electrophoresis (a
100 bp molecular marker ladder
was used).
Note: The rbcl2f and RBCL-fonfana were the primers used to amplify the RBCL gene during real-time PCR. |
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