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Determination of Water Use Efficiency for Arabidopsis thaliana
拟南芥水分利用效率的测定   

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Abstract

Water use efficiency (WUE) is a quantitative measurement of how much biomass or yield is produced per amount of water used. It is an important physiological factor for agriculture, especially in areas with a limited accessibility of water. It is also crucial in a better understanding of drought tolerance and drought resistance. The most common method to measure the WUE of individual plants is to weigh each pot or container to monitor the water loss and to harvest and measure biomass or yield at the end of the experiment. Since water can be lost from the soil surface through evaporation, there is a need to perform WUE measurements in a closed system. Here, we describe a simple method for WUE determination for Arabidopsis thaliana.

Keywords: Water use efficiency (WUE)(水分利用效率(WUE)), Arabidopsis thaliana(拟南芥), Biomass(生物量), Plants(植物), Falcon tubes(猎鹰管)

Materials and Reagents

  1. Arabidopsis seeds
  2. Soil
  3. Perlite
  4. Tap water
  5. Aluminium foil

Equipment

  1. 50 ml Falcon tubes
  2. Tweezers
  3. Burner
  4. Milligram balance with a resolution of minimum 3 decimal place (0.001 g) readability
  5. Transparent, tight container with a cover
  6. Cold room/fridge
  7. Growing chamber/phytotron
  8. Incubator (with a possibility to set temperature 105 °C or above)

Procedure

  1. In a container, prepare a mixture of soil and perlite in a 1:1 proportion.
  2. Fill in 50 ml Falcon tubes with soil-perlite mixture to the 70-80% of Falcon volume.
  3. To each Falcon pour out 35 ml of water and wait till the water sinks into soil-perlite mixture. 35 ml is normally sufficient to grow plants to the stage of 4-week-old.
  4. In a separate container, moisten soil with water in a 1:1 ratio.
  5. Full the Falcon tubes to the brim with well-moistened soil.
  6. Using a burner, heat the tweezers and use them to make a hole (1.5 to 2 mm wide) in a central part of the Falcon tube cap. Another hole can be made closer to the cap edge (see Figure 2) in order to supplement the system with water (in case of growing plants longer than 4 weeks).
    Note: While making a whole, move the tweezers’ end from the upper side of a cap towards the bottom side to avoid protrusion of plastic scraps that could wound the seedling.
  7. Turn off each brim-filled Falcon tube with the cap.
  8. Wrap Falcon tube with aluminium foil or other non-transparent material to avoid algae growth.
  9. In the central hole place 5 to 10 Arabidopsis seeds.
  10. Store falcon tubes in 4 °C in a humid environment (for example in a closed transparent container with a water-covered bottom) for two days.
  11. After two days, move the whole system (Falcon tubes placed in a transparent water-containing container) to the growing chamber/phytotron.
  12. After plants germinate (after about 5-7 days), remove excessive seedlings (using tweezers), leaving only one seedling in a hole (Figure 1).


    Figure 1. Falcon tubes with single seedling each – ready to determine Falcon weight at the beginning of experiment (W0)

  13. Weigh each falcon tube and write down the falcon weight (g) at the beginning of experiment (W0).
  14. Grow plants for 4 weeks in a growing chamber/phytotron.
  15. After 4 weeks (Figure 2), decapitate the rosettes and dry them for 3 h at 105 °C. Weigh dry mass for each rosette (mg) (DW).


    Figure 2. Rosettes after 4 weeks of experiment – ready for decapitation

  16. After removal of rosette, weigh each falcon tube and write down the falcon weight (g) at the end of the experiment (W).
  17. Calculate the water loss: W0 (g) – W (g) = Water used (g). For water 1 g = 1 ml.
  18. Calculate dry weight per unit of water used (mg DW x /ml of water used).

Acknowledgments

Please also cite the following publication when citing this protocol: Wituszyńska et al. (2013). This work was supported by the Welcome/2008/1 Program operated within the framework of the Foundation for Polish Science and co-financed by the European Regional Development Fund.

References

  1. Wituszynska, W., Slesak, I., Vanderauwera, S., Szechynska-Hebda, M., Kornas, A., Van Der Kelen, K., Muhlenbock, P., Karpinska, B., Mackowski, S., Van Breusegem, F. and Karpinski, S. (2013). Lesion simulating disease1, enhanced disease susceptibility1, and phytoalexin deficient4 conditionally regulate cellular signaling homeostasis, photosynthesis, water use efficiency, and seed yield in Arabidopsis. Plant Physiol 161(4): 1795-1805.

简介

水利用效率(WUE)是每使用的水的量产生多少生物量或产量的定量测量。 它是农业的一个重要生理因素,特别是在水的可及性有限的地区。 这对于更好地了解耐旱性和抗旱性也至关重要。 测量单个植物的WUE的最常见方法是称重每个盆或容器以监测水损失并在实验结束时收获和测量生物量或产量。 由于水可能通过蒸发从土壤表面损失,因此需要在封闭系统中进行WUE测量。 在这里,我们描述了拟南芥的WUE测定的简单方法。

关键字:水分利用效率(WUE), 拟南芥, 生物量, 植物, 猎鹰管

材料和试剂

  1. 拟南芥种子
  2. 土壤
  3. 珍珠岩
  4. 自来水
  5. 铝箔

设备

  1. 50ml Falcon管
  2. 镊子
  3. 燃烧器
  4. 毫克平衡,分辨率至少为小数点后三位(0.001克)的可读性
  5. 带有盖子的透明,紧凑容器
  6. 冷室/冰箱
  7. 生长室/phytotron
  8. 孵化器(可能设置温度105°C或以上)

程序

  1. 在容器中,以1:1的比例制备土壤和珍珠岩的混合物。
  2. 在土壤 - 珍珠岩混合物中填充50毫升Falcon管70-80%的Falcon体积。
  3. 向每个Falcon倒出35毫升的水,等待水沉入土壤 - 珍珠岩混合物中。 35毫升通常足以使植物生长到4周龄的阶段
  4. 在单独的容器中,用水以1:1的比例湿润土壤
  5. 将猎鹰管充满边缘,并用湿润的土壤
  6. 使用燃烧器,加热镊子,并使用它们在Falcon管帽的中心部分做一个孔(1.5到2毫米宽)。可以使另一个孔更靠近盖边缘(见图2),以便用水补充系统(在生长超过4周的植物的情况下)。
    注意:在做整体的时候,将镊子的端部从帽子的上侧移向底侧,以避免塑料碎屑突出,从而可能伤害幼苗。
  7. 用盖子关闭每个装满边缘的猎鹰管。
  8. 用铝箔或其他不透明材料包裹Falcon管,以避免藻类生长
  9. 在中心孔中,5至10个拟南芥种子
  10. 在4℃的潮湿环境(例如在有水的底部的封闭透明容器中)存放猎鹰管2天。
  11. 两天后,将整个系统(放置在透明含水容器中的Falcon管)移动到生长室/phytotron。
  12. 植物发芽后(约5-7天),除去过量的幼苗(使用镊子),在孔中只留下一个幼苗(图1)。


    图1. Falcon管单个幼苗 - 准备在实验开始时确定Falcon的重量(W0)

  13. 在实验开始时(W0),称重每个猎鹰管并记下猎鹰重量(g)。
  14. 在生长室/phytotron中种植植物4周。
  15. 4周后(图2),将玫瑰花瓣断头并在105℃下干燥3小时。 每个玫瑰花的干重(mg)(DW)。


    图2.经过4周的实验 - 已准备好斩首后的玫瑰花。

  16. 取出玫瑰花后,称重每个猎鹰管,并写下实验结束时的猎鹰重量(g)(W)。
  17. 计算水损失:W0(g)-W(g)=使用的水(g)。 对于水1g = 1ml。
  18. 计算每单位所用水的干重(mg DW x/ml使用的水)

致谢

请引用此协议时引用以下出版物:Wituszyńska(2013)。这项工作得到了在波兰科学基金会框架内开展并由欧洲区域发展基金共同资助的欢迎/2008/1计划的支持。

参考文献

  1. Wituszynska,W.,Slesak,I.,Vanderauwera,S.,Szechynska-Hebda,M.,Kornas,A.,Van Der Kelen,K.,Muhlenbock,P.,Karpinska,B.,Mackowski, Breusegem,F。和Karpinski,S。(2013)。 病变模拟疾病1,增强的疾病易感性1和植物抗毒素缺陷4有条件地调节细胞信号稳态,光合作用,水分利用效率和种子产量。植物生理 161(4):1795-1805。
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Copyright: © 2014 The Authors; exclusive licensee Bio-protocol LLC.
引用: Readers should cite both the Bio-protocol article and the original research article where this protocol was used:
  1. Wituszynska, W. and Karpiński, S. (2014). Determination of Water Use Efficiency for Arabidopsis thaliana. Bio-protocol 4(3): e1041. DOI: 10.21769/BioProtoc.1041.
  2. Wituszynska, W., Slesak, I., Vanderauwera, S., Szechynska-Hebda, M., Kornas, A., Van Der Kelen, K., Muhlenbock, P., Karpinska, B., Mackowski, S., Van Breusegem, F. and Karpinski, S. (2013). Lesion simulating disease1, enhanced disease susceptibility1, and phytoalexin deficient4 conditionally regulate cellular signaling homeostasis, photosynthesis, water use efficiency, and seed yield in Arabidopsis. Plant Physiol 161(4): 1795-1805.
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Michael Itam
Tottori University, Japan
Hello thank you for your article. I want to carry out drought stress experiment on Arabidopsis by withholding water. Please guide me.

Regards,
Michael
7/12/2016 12:04:09 AM Reply