03-Mineral Nutrition Spring 2011.pdf

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Biology 3460 – Weeks 2, 4, 6, & 8 1

Biology 3460 - Plant Physiology - Lab Exercise 3Plant Mineral Nutrition

Objectives:This lab exercise is intended to:

(1) introduce students to the mineral nutrient requirements of plants,(2) illustrate sand culture as a technique to grow plants under controlled nutrient

conditions,(3) provide experience making plant nutrient solutions and calculating nutrient

concentrations in different solutions,(4) test of the effect of different sources of nitrogen on growth of grasses.

IntroductionMineral nutrition is the study of how plants obtain and use basic mineral nutrientsnormally taken up by roots from the soil solution. Essential mineral nutrients required by

plants are shown in Table 5.1 in your textbook (attached). Access to these nutrients insufficient supply, along with adequate water and sunlight allows plants to synthesize allthe other components they require for growth. An empirical formulation of a completenutrient solution for plants was developed by Dennis Hoagland, a researcher inCalifornia, USA. There have been several modifications of Hoagland’s solution over theyears with a typical example of a “modified Hoagland’s solution” illustrated in Table 5.3of your textbook (attached). Hoagland’s solutions typically provide nutrient elem ents inratios related to the nutrient requirements by plants, but the nutrient concentrations in fullstrength modified Hoagland’s solution can be several times higher than a plant wouldexperience in the natural world. When growing non-agricultural plants it is oftennecessary to dilute the modified Hoagland’s solution in order to prevent injury due totoxicity or salinity stress, particularly with young seedlings.

Typical modified Hoagland’s solutions provide nitrogen as both ammonium (NH 4+) and

nitrate (NO 3-) ions. Most plants grow best when nitrogen is supplied from both

ammonium and nitrate. If nitrogen is supplied as only one type of ion (either nitrate orammonium) large changes in the pH of the nutrient solution can occur and this can thendisrupt the uptake of other required nutrients such as calcium, magnesium or iron. Plantsgenerally grow better when they have access to both ammonium and nitrate because this

promotes cation-anion nutrient balances within the plant.

Instructions for making the macro- nutrient stock solutions for modified Hoagland’ssolution are shown below. Three different formulations are illustrated, one with bothammonium and nitrate supplied (typical Hoagland’s solution) and then two solutions inwhich nitrogen only in the form of either nitrate or ammonium is supplied. In this labexperiment, plants will be grown in one of three treatments (both ammonium and nitrate,nitrate only, and ammonium only) to test the effect of nitrogen source on plant growth(biomass (dry weight) production). Note that it is not possible to keep the concentrationsof all macro-nutrients at identical concentration in all three versions of the modifiedHoagland’s solution (Calcium concentration is modified in the nitrate -only solution andSulphur concentration is modified in the ammonium-only solution).



Biology 3460 – Weeks 2, 4, 6, & 8 2

In addition to the macro-nutrients shown in the nutrient solution recipes illustrated below,the modified Hoagland’s solution also provides micro -nutrients from a prepared stocksolution and iron from a chelated-iron solution, which keeps the iron in a soluble formand prevents it from precipitating out of solution and making it unavailable to the plants.

Part A. The Nutrient Solutions

Modified Hoagland’s Solution (both NO 3 and NH 4 provided)

Macro-nutrient Stock Solutions:

1M Calcium nitrate [Ca(NO 3)2 4H 2O]1M Potassium nitrate [KNO 3]1M Magnesium sulphate [MgSO 4 7H 2O]1M Ammonium phosphate [NH 4 H2 PO 4]

To make 4 litres, add: 4 ml calcium nitrate 6 ml potassium nitrate 2 ml magnesium sulphate 6 ml ammonium phosphate 2 ml iron + 2 ml micronutrients

Top up to 4 L with distilled water

Modified Nutrient Solution (only NO 3 provided)

Macro-nutrient Stock Solutions: 1M Calcium nitrate [Ca(NO 3)2 4H 2O]1M Magnesium sulphate [MgSO 4 7H 2O]1M Potassium phosphate (monobasic) [KH 2PO 4]

To make 4 litres, add: 10 ml calcium nitrate 2 ml magnesium sulphate 6 ml potassium phosphate 2 ml iron + 2 ml micronutrients




Biology 3460 – Weeks 2, 4, 6, & 8 3

Modified Nutrient Solution (only NH 4 provided)

Macro-nutrient Stock Solutions:

1M Calcium chloride [CaCl 2 2H 2O]1M Potassium phosphate (monobasic) [KH 2PO 4]

1M Magnesium sulphate [MgSO 4 7H 2O]1M Ammonium sulphate [(NH 4)2SO 4]

To make 4 litres, add: 4 ml calcium chloride 6 ml potassium phosphate 2 ml magnesium sulphate 10 ml ammonium sulphate 2 ml iron + 2 ml micronutrients


Protocol :Work as individuals to complete this part of the lab.

1. Calculate the concentration (mmol/L) of each of the macro-nutrients for the threemodifie d Hoagland’s solutions and fill in the table below. A sample calculation is

provided.

Sample Calculation:For the Hoagland’s solution with both ammonium and nitrate present, the concentration of

potassium (K+) is determined as follows:

6 ml of 1 M potassium nitrate was added to a bottle and the volume is made up to 4 L.

6 ml x 1 mmol/ml (1 M) potassium nitrate = 6 mmol of potassium nitrate

6 mmol potassium nitrate in 4 L of solution is: 6 mmol/4 L = 1.5 mmol/L

Table 1. Concentration of individual macronutrients available in three modified Hoagland’ssolutions.Macro-Nutrient Both NO 3

- & NH 4+

(mmol/L)NO 3

- only(mmol/L)

NH 4+ only

(mmol/L) N

P

K

Ca

Mg

S



Biology 3460 – Weeks 2, 4, 6, & 8 4

Part B. The Experimental Design

Plants will be grown in sand culture in each of the three nutrient treatments, under the samelight conditions. We will measure total dry biomass of the plants at the end of a four-weekgrowing period.

Protocol:Work as a group of three students to complete this part of the lab.

1. Send one of your group members to help with mixing the first batches of the threenutrient solutions, according to the recipes outlined in Part A.

2. Prepare one plant pot for each of the three treatments (both NO 3 and NH 4, only NO 3, and only NH 4). Be sure to label the pots with the treatment type and a groupidentifier. Add a layer of polyfilter wool to the bottom of the pot to prevent sandfrom escaping through the holes. Fill the pot with sand up to about 2 cm below thelip of the pot.

3. Several grass seedlings (your instructor will inform of the species used) have beengerminated in Petri dishes in Promix soil mixture. Carefully choose four seedlingsfor each treatment (will need 12 seedlings in total), be sure to choose healthy-looking seedlings of about the same stage of germination. Remove the seedlingsvery gently from the Petri dish, and rinse them under a gentle stream of water toremove as much soil from the roots as possible.

4. Moisten the sand in the prepared plant pots with the appropriate nutrient solution,then make wells in which to plant the four seedlings for each treatment. Poke awell with your finger deep enough to support the roots of the seedling, but not sodeep that the shoot will be under sand once planted. Place the seedling into the wellthen gently push sand around it to fill in the well and support the seedling. Be sureto space the four seedlings evenly throughout the pot.

5. Once you have completed a pot for each treatment, place them in the appropriatelylabeled plastic tub. Your lab instructor will fill the tubs with the nutrient solutions

prepared in Step 1. Nutrient solutions will be changed twice a week. You may beasked in future labs to mix fresh nutrient solutions and refill the tubs.

6. At the end of the four-week growing period, cut the plants off the roots right at thesurface of the sand, place them in an envelope for each treatment (label your groupnames and the treatment type on the envelope). The above-ground plant materialwill be dried in a drying oven for two weeks. In the next lab period we will collectclass data for the above-ground biomass for each treatment.



Biology 3460 – Weeks 2, 4, 6, & 8 5

Treatment Above-ground Biomass (g)Bench 1 Bench 2 Bench 3 Bench 4 Bench 5 Mean

SDBoth NO 3

-

NH 4+

NO 3

- only

NH 4+ only

Analysis :1. Use class results to prepare a figure that shows the effect of mineral nutrition treatment

on the above-ground biomass of the plants. Mean biomass values with standarddeviations should be plotted.

2. Using a Kruskal Wallis Test, determine if there is a statistically significant difference between the treatments.

3. What effect did the different nutrient regimes have on plant growth? Why? Explain.



Biology 3460 – Weeks 2, 4, 6, & 8 6



Biology 3460 – Weeks 2, 4, 6, & 8 7

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03-Mineral Nutrition Spring 2011.pdf