Planting and Harvesting Procedures for Bio 105 Two-Factor Experiments
  1. Design your experiment. (See previous Bio 105 student greenhouse projects)
  2. Things to keep in mind:
    1. The main purpose of these experiments is to examine the INTERACTION between two factors (e.g. "do you expect the effect of planting date to have a greater effect at high nutrient or low nutrient? ..."why?")
    2. Factors that work:
        - seed orientation (root up vs down)(2 plants/cup: one up, one down)
        - planting date (2 plants/cup: one early, one several days later)
        - herbivory (early & lots e.g. all except one leaf)(2 plants/cup: one clipped one not clipped)
        - nutrients
        - shade (limit height of shade tent & get on early)
    3. Factors that don't work:
        - planting depth
        - water-logging
        - compaction
        - soil temperature
        - wind
  3. Randomization.
    Determine the treatments (e.g. High herbivory/High nutrient; High herbivory/Low nutrient; Low herbivory/High nutrient; Low herbivory/Low nutrient). If you have 4 treatments and 60 plants total, you will have 15 plants in each treatment. Random number tables are available at the greenhouse. The first 15 numbers will be assigned to plants in Treatment 1. The next 15 numbers will be assigned to plants in Treatment 2, and so on. Note: For those using light intensity as a factor, you have to do a semi-random set-up and will use a random number table that randomizes 1-30 seperately from 31-60. This is because all the shade plants must go together in one tray and all the high light plants go together in the other tray. Each treatment will have 15 plants, but numbers 1-30 will be randomly assigned to the high light plants, and numbers 31-60 will be randomly assigned to the low light plants.
  4. Prepare your cups:
    1. Use a spatula to punch six drainage holes in the bottoms of your Styrofoam cups. It is important to be consistent in your pattern so that all cups drain the same way.
    2. Use your Table of Random Numbers to label your cups using a BLACK Sharpie Permanent marker. Non-permanent markers run and Sharpies of other colors fade quickly in the greenhouse.
    3. Label the cups: (a) random number (b) treatment (c) your group number.
    4. Mark the rim of the cup to designate the different treatments. This makes it easier to see which cups get which treatments without having to pick each cup up out of the tray to read the label.
    5. Stack the cups in NUMERICAL ORDER.
  5. Fill your cups with sand in NUMERICAL ORDER. This RANDOMIZES both the sand and the filling of the cups so that one treatment isn’t biased over the others. Sand should be 1/2 inch from top of the cup.
  6. Place cups in the trays in a honeycomb pattern so that there are 5 cups per row and six rows in each tray. (the first tray will have cups 1-30, the second tray will have cups 31-60). The honeycomb pattern keeps the cups from falling over in the trays. The cups go into the trays in NUMERICAL ORDER, from left to right across the tray. There are two reasons for this:
    1. The plants from each treatment are randomly distributed throughout your trays, thus randomizing any environmental variability.
    2. When cups are removed from the tray (for taking measurements), you will know exactly where to put them back when you are done.
  7. When planting Abutilon, use ONLY the seeds that have split open and have begun to grow. Plant the seeds with the emerging root DOWN. Seeds should be planted 1/4 to 1/2 inch deep. Plant the cups in NUMERICAL ORDER (insures seed size is distributed among all your treatments). Plant twice as many seeds as you want to have growing in your cups (e.g. if your design calls for 2 seeds/ cup..then plant 4 and we will thin any "extras"....this is to ensure that you don't end up with "empty cups").
  8. Fill out the form (This will go in a ziploc bag and stay with your trays.)
    1. Group Letter.
    2. Both factors for your experiment.
    3. How many plants you want per cup after they have been thinned.
    4. Names and email addresses for everyone in your group.
  9. Once you are finished planting, the greenhouse staff or T.A.s will show you where your trays are supposed to go on the benches. Greenhouse staff will water your plants.
  10. This is just the beginning! Be thinking about possible interactions and why interactions are important.
  11. Data Collection and Harvesting:
    1. Determine what kinds of data you need to collect in order to determine if there is an interaction. (e.g. if one seed is planted early and it starts growing before the other seed in the same cup... will it have a greater advantage at high nutrient or low nutrient? i.e. if you are looking at planting date or seed orientation ...then be sure to record day of seed emergence).
    2. Check your plants weekly and take 'non-destructive' measurements. (e.g. height, size of leaves, stem diameter)
    3. Plants will be harvested approximately 6 weeks after planting.
    4. Before you harvest your plants: line them up by treatment and take a photo of them (greenhouse has digital camera). You'll then be able to determine if mislabelling or errors occurred during data collection or analysis.
    5. What to measure when harvesting: height, stem diameter, above ground biomass, root biomass (wash the sand off well...sand is heavier than roots!)
    Revised August 2003