Measurement of Net High-affinity Urea Uptake in Maize Plants
玉米植株高亲和力转运系统吸收尿素的测定方法
发布: 2015年06月05日第5卷第11期 DOI: 10.21769/BioProtoc.1490 浏览次数: 9470
评审: Samik BhattacharyaSriema L. WalawageAnonymous reviewer(s)
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参见作者原研究论文
The authors used this protocol in:
Sep 2014
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Abstract
Despite its extensive use as a nitrogen fertilizer, the role of urea as a directly accessible nitrogen source for crop plants is still poorly understood. So far, the physiological and molecular aspects of urea acquisition have been investigated only in a few plant species highlighting the importance of a urea transporter in roots, DUR3 (Kojima et al., 2007; Wang et al., 2012; Zanin et al., 2014a). Regarding maize plants, a crop that needs a large amount of urea fertilizer, the capability to take up urea via an inducible and high-affinity transport system has been recently characterized (Zanin et al., 2014a; Zanin et al., 2014b). Here, we described a small-scale protocol suitable for the measurement of urea net high-affinity uptake in roots of intact maize plants.
Keywords: Transport (运输)
Nitrogen (氮)
Root (根)
Corn (玉米)
Method (方法)
Materials and Reagents
- Maize seeds (Zea mays L., cv. PR33T56, Pioneer Hi-bred Italia S.p.A., Parma, Italy)
- Potassium hydroxide (KOH) (Sigma-Aldrich, catalog number: 60370 )
- Calcium sulphate (CaSO4) (Sigma-Aldrich, catalog number: 12090 )
- Urea [CO(NH2)2] (Sigma-Aldrich, catalog number: 15604 )
- Diacetylmonoxime [CH3C(=NOH)COCH3] (Sigma-Aldrich, catalog number: 31550 )
- Thiosemicarbazide (NH2CSNHNH2) (Sigma-Aldrich, catalog number: 89050 )
- Sulphuric acid (H2SO4) (Sigma-Aldrich, catalog number: 320501 )
- Ortho-phosphoric acid (H3PO4) (Sigma-Aldrich, catalog number: W290017 )
- Ferric chloride hexahydrate (FeCl3) (Sigma-Aldrich, catalog number: 12319 )
- Sterile deionized water
- KCl (Sigma-Aldrich, catalog number: P9541 )
- CaSO4 (Sigma-Aldrich, catalog number: 12090 )
- MgSO4 (Sigma-Aldrich, catalog number: 746452 )
- KH2PO4 (Sigma-Aldrich, catalog number: P9791 )
- NaFe-EDTA (Sigma-Aldrich, catalog number: 03650 )
- H3BO3 (Sigma-Aldrich, catalog number: B7901 )
- MnSO4 (Sigma-Aldrich, catalog number: 221287 )
- ZnSO4 (Sigma-Aldrich, catalog number: Z1001 )
- CuSO4 (Sigma-Aldrich, catalog number: C3036 )
- Na2MoO4 (Sigma-Aldrich, catalog number: M1651 )
- Nutrient solution (see Recipes)
- Urea solution stock (see Recipes)
- Mixed colour reagent (see Recipes)
- Mixed acid reagent (see Recipes)
- Colour development reagent (see Recipes)
Equipment
- Growth chamber and hydroponic growing system (including plastic net, plastic pots)
- pH meter (Jenway, model: 3510 )
- Plastic box (15 x 10 cm; H 4 cm; Figure 1A, alternatively you can use the bottom of the pipette tips, Sigma-Aldrich, catalog number: P5161 )
- 0.2 ml 96-well plate (AB ANALITICA Advanced Biomedicine, catalog number: B50601 ; Figure 1B)
- Clear 96-well microplate with flat bottoms (STARLAB, catalog number: S1837-9600 ; Figure 1C)
- Sealing tapes, optically clear (SARSTEDT AG, catalog number: 95.1994 )
- Thermocycler (Eppendorf, model: Mastercycler® personal )
- Orbital shaker (Janke & Kunkel IKA-Labortechnik, model: KS 501D )
- Spectrophotometric multiwell plate reader (TECAN, model: GENios Microplate Reader )
- Timer
- Pipettes (Eppendorf, model: 0.5-10 μl, 20-200 μl, 100-1,000 μl, Eppendorf Reference® 2 ) and tips
- 1.5 ml plastic tubes (Eppendorf, catalog number: 0030 125.150 )
- Absorbent paper (Sigma-Aldrich, catalog number: Z270849 )
Figure 1. Plastic equipment. A. plastic box; B. 0.2 ml 96-well plate; C. Clear 96-well microplate.
Procedure
文章信息
版权信息
© 2015 The Authors; exclusive licensee Bio-protocol LLC.
如何引用
Zanin, L., Tomasi, N. and Pinton, R. (2015). Measurement of Net High-affinity Urea Uptake in Maize Plants. Bio-protocol 5(11): e1490. DOI: 10.21769/BioProtoc.1490.
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分类
植物科学 > 植物生理学 > 营养
植物科学 > 植物新陈代谢 > 氮
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