The authors would like to thank California Department of Food and Agriculture for funding of the project which led to the development of this protocol. CDFA SCB11019.

1. Dikim <ol><li> Fiş tepsileri doldurun (128 hücre; 3 cm kare hücreleri ile 28 x 54 cm ve 5 cm derinliğinde) fiş toprak karışımı ile. Hücrelerin düzgün dolum yardımcı olmak için boş bir fiş tepsisini kullanarak her tepsiye toprak sıkıştırmak. </li><li> Bitki marul tohumu ¼ derinlemesine hücre başına 2-3 tohum inç. Bitki Çoğaltılmış tepsiler tüm deneysel hatları kuraklık hem stresli ve kuraklık deneme için kontrol tepsiler sağlamak. </li><li> Deliksiz bir taban tepsisine fiş tepsilerini yerleştirin. </li><li> Ters tepsiye veya plastik kubbe Su tepsiler ve kapak. Çimlenme normal olarak 48-72 saat içinde, gerçekleşir. Çimlenme sonra kapağı çıkarın ve sera içine tepsileri taşımak. NOT: Bu protokol, aynı zamanda büyüme odalarına gerçekleştirilebilir. </li></ol> 2. Büyüme ve Bitki Bakımı <ol><li> Çimlenme sonrası 1-4 hafta gerektiğinde su tepsiler, normalde bu toprak alt tepsiyi doldurarak ve izin vererek, 2-3 kez haftalık uzun süre oturmak (~ 1 saat) ışınları doymuş hale içinBu gibi suda s olumsuz bitki sağlığı ve kök gelişimini etkileyebilecek ve kuraklık ekranı etkileyebilir. </li><li> 1.5 tk / galon bir oranda karıştırılmış ticari çözünür 20-20-20 gübresi ile haftada bir kez tepsileri döller. </li><li> Hücre 1-2 hafta başına bir bitki İnce tepsiler çimlenme sonrası. </li></ol> 3. başlatılıyor Kuraklık Stres <ol><li> Bitkiler 2 gruba dört haftalık ayrı tepsiler zaman. Şiddetli su stresine tabi bir grup. Yanı sıra kontrolleri sulanan diğer tepsileri kullanın. Deney boyunca adım 2.1 gibi kontrol grubu sulayın. Kuraklık stresi başlangıcında deney kalanı için tüm tepsileri gübreleme askıya alma. </li><li> Deneysel kuraklık dönemi bir hafta içinde oluşur. Kuraklık stresi deneme (Gün 0) ilk gününde su ile düşük tepsileri doldurmak ve fiş tepsilere toprak sonra tam doymuş hale kasetlerden suyu tahliye için izin verir. T başlamasından hemen önce toprak doyurulmasıHe stres dönemi fiş tepsiler bireysel hücreler arasında toprak nem herhangi bir varyasyonu en aza indirmek için yardımcı olacaktır. </li></ol> Parametreler ile Temel Kuraklık Stres Ekranı yapılması 4. Ölçülen edilecek NOT: Aşağıdaki adımlar Tablo 1 kuraklığa stres deneme süresi her gün alınan ölçümlerin bir program sağlarken kuraklık ekran boyunca kaydedilecektir üç fizyolojik ölçümler toplamak için nasıl özetlemektedir. <ol><li> Gün 0, 2, deneme 4 Yaprak Oransal Su İçeriği (OSK) ölçün. <ol><li> Tepsiden bir marul bitki Klip ve 2 yaprak çıkarın. </li><li> Bir 9. mantar delici (~ 1.6 cm çapında) kullanarak her yapraktan 3 diskleri Punch. Bir örnek olarak yaprak yumruklar birleştirin. </li><li> Yaprak diskleri tartılır, bu RWC hesaplanmasında kullanılan taze ağırlığı (FW), bir değerdir, daha sonra da bir petri plakasında diskleri yerleştirin. </li><li> Tüm plaka için yeterli Damıtılmış su ekleyinow tüm yaprak diskleri float. Bu tamamen RWC analiz için şiş ağırlıkları toplamak için yaprak diskleri hidrat etmektir. Yaprak diskleri, oda sıcaklığında 24 saat boyunca kalmasına izin verin. </li><li> Plakalardan artık tamamen hidrate yaprak diskleri çıkarın ve yavaşça diskleri tartı önce kağıt havlu ile yaprak diskleri dış kurulayın. RWC hesaplama için abartılı ağırlık (TW) olarak bu ağırlıkları kaydedin. </li><li> Bir kağıt laboratuarda yerleştirin yaprağı diskleri, 24 saat boyunca 55 ° C&#39;de inkübatör içinde silme ya da açık bir petri filtre kağıdı diski ve kurutulur. NOT: Kağıt kurutma sırasında petri yapışmasını yaprak diskleri engeller. </li><li> Kuru yaprak diskleri ve RWC hesaplama için rekor olarak kuru ağırlıkları (DW) tartılır. </li><li> Smart ve Bingham 11,12 tarafından tarif edildiği gibi OSK Weatherley formülü kullanılarak hesaplayın. OSK = (FW-DW / TW-DW) x 100 FW taze ağırlık, TW = şiş ağırlık, DW = kuru ağırlığı = </li><li> Deneme her germplazma ve tedavi için yineleyin 4.1. </li></ol></li><li> Kaydedin bitki solgunluk kuraklık stresi deneme gün 1 ve bitkilerin% 100 normal günde 4 ya da 5, soldu zaman biten günde bir kez bir başlangıç. </li><li> Kurtarma fazı ve büyüme diferansiyel <ol><li> 6. günde, kuraklığa stres dönemi sonuçlandırmak ve stresli bitkiler deneme toparlanma sürecine girmek. Kontrol tepsiler gibi, 2.1 adım, su ile düşük tepsiler doldurun ve kuraklık vurguladı tepsiler standart sulama takvimi başlamadan önce 24 saat bekletin. </li><li> Tüm bitkileri, 10 gün süre ile iyileşmeye izin verin. </li><li> 10 gün post kuraklığa stres, her bitkinin toprak bölümü tüm yukarıdaki hasat ve taze ağırlığı kaydetmek. </li></ol></li></ol>

<ol>
	<li>Dijk, A. I. J. M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Millennium+Drought+in+southeast+Australia+(2001&#8211;2009):+Natural+and+human+causes+and+implications+for+water+resources,+ecosystems,+economy,+and+society.">The Millennium Drought in southeast Australia (2001&#8211;2009): Natural and human causes and implications for water resources, ecosystems, economy, and society.</a> Water Resour. Res. 49, (2013).</li><li>Aghakouchak, A., Feldman, D., Stewardson, M. J., Saphores, J., Grant, S., Sanders, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Australia&#8217;s+Drought:+Lessons+for+California.">Australia&#8217;s Drought: Lessons for California.</a> Science. 343, 1430 (2014).</li><li>Tolomeo, V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=California+Agricultural+Statistics+2012+Crop+Year.">California Agricultural Statistics 2012 Crop Year.</a> USDA National Agricultural Statistics Service Pacific Regional Office-California. , (2013).</li><li>Cavagnaro, T., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Climate+Change:+Challenges+and+Solutions+for+California+Agricultural+Landscapes.+White+paper+CEC-500-2005-189-SF.">Climate Change: Challenges and Solutions for California Agricultural Landscapes. White paper CEC-500-2005-189-SF.</a> California Climate Change Center. , (2005).</li><li>Lobell, D. B., Gourdji, S. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Influence+of+Climate+Change+on+Global+Crop+Productivity.">The Influence of Climate Change on Global Crop Productivity.</a> Plant Physiol. 160, 1686-1697 (2012).</li><li>Jackson, L. E., Stivers, L. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Root+distribution+of+lettuce+under+commercial+production:+implications+for+crop+uptake+of+nitrogen.">Root distribution of lettuce under commercial production: implications for crop uptake of nitrogen.</a> Biological Agriculture and Horticulture. 9, 273-293 (1993).</li><li>Jackson, L. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Root+architecture+in+cultivated+and+wild+lettuce+(Lactuca+spp).">Root architecture in cultivated and wild lettuce (Lactuca spp).</a> Plant. Cell and Environment. 18, 885-897 (1995).</li><li>Malcom, S., Marshall, E., Aillery, M., Heisey, P., Livingston, M., Day-Rubenstein, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Agricultural+Adaptation+to+a+Changing+Climate:+Economic+and+Environmental+Implications+Vary+by+U.S+Region.+USDA+Economic+Research+Service.">Agricultural Adaptation to a Changing Climate: Economic and Environmental Implications Vary by U.S Region. USDA Economic Research Service.</a> Economic Research Report Number. 136, (2012).</li><li>Chaves, M. M., Maroco, J. P., Pereira, J. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Understanding+plant+responses+to+drought-+from+genes+to+the+whole+plant.">Understanding plant responses to drought- from genes to the whole plant.</a> Funct. Plant Biol. 30, 239-264 (2003).</li><li>Ingram, J., Bartels, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Molecular+Basis+of+Dehydration+Tolerance+in+Plants.">The Molecular Basis of Dehydration Tolerance in Plants.</a> Annu. Rev. Plant Physiol. Plant Mol. Biol. 47, 377-403 (1996).</li><li>Weatherley, P. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Studies+in+the+water+relations+of+the+cotton+plant.+I.+The+field+measurement+of+water+deficits+in+leaves.">Studies in the water relations of the cotton plant. I. The field measurement of water deficits in leaves.</a> New Phlytol. 49, 81-97 (1950).</li><li>Smart, R. E., Bingham, G. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Rapid+Estimates+of+Relative+Water+Content.">Rapid Estimates of Relative Water Content.</a> Plant Physiol. 53, 258-260 (1974).</li></ol>

The authors declare that they have no competing financial interests.

Ekran için örnek sayısının Hususlar. gerekli numune sayısı bu ekrandan verilerin istenen kullanıma bağlı olmalıdır. Yayın kalitesi sonuçları istenirse her satırından 3 ayrı bitkiler (3 biyolojik çoğaltır) hasat ve kalite istatistiksel analiz için yeterli puan vermek için 2 deneysel çoğaltır en az gerçekleştirmek için tavsiye edilir. İstenilen sonuç ise sadece hızlı daha sıkı veya karmaşık su stres deneyleri, daha az örnekleri gerçekleş...

Water availability for irrigation has been a concern across much of the United States and globally for decades, but research into the response to drought-stress, along with other abiotic stresses, has lagged behind work in the areas of disease and insect resistance, at the industrial, academic, and governmental levels largely due to a lack of funding. Water availability for agriculture has historically been only an afterthought at the level of policy makers. Recently, due to several severe droughts in important agricultural production regions in both the United States and Australia1,2 fresh water availability has been thrust into the spotlight at both the national and international levels leading to many more resources being directed towards research into developing drought-tolerant cultivars of the major grain crops. While this shifting focus toward the development of drought-tolerant major crops is beneficial, like many areas of plant research specialty crops have largely been left behind. 
A severe drought is currently limiting vegetable production in California, the largest production region for Lettuce (Lactuca sativa L.) in the United States3. These short-term weather patterns coupled with legislative and judicial action along with long-term climate change4,5 have combined to reduce water available for agriculture in many of the most productive regions of California. Lettuce production in California represents a 1.5 billion dollar industry accounting for nearly 80% of lettuce production in the United States3. Leafy vegetables have high leaf water content and lettuce, in particular, has a shallow root system6,7 which leaves the crop vulnerable to water-stress. In lettuce, as in all crops the development of drought-tolerant varieties will become increasingly important as fresh water supplies for irrigation become more constrained8.
This protocol lays out a method by which an initial screen was performed on a large collection of lettuce germplasm in order to identify a pool of potentially drought-tolerant candidate lines for use in physiological, molecular, and genetic studies to identify specific drought-tolerance traits. These candidates can also be used directly as sources of drought-tolerance in breeding programs to improve water use efficiency in commercial cultivars. This protocol was developed specifically to address the challenges that arise during the course of any screen of an extensive germplasm collection, especially issues of space availability and labor. Also, the protocol as presented was developed for use in lettuce, but has been successfully adapted for use in the screening of a germplasm collection of spinach (Spinacia oleracea) for potential drought-tolerance and could be modified simply to screen any leafy vegetable crop. 
An important consideration before initiating a drought-tolerance screen is to understand what this method is and is not. This protocol is meant to represent a rapid method by which a large germplasm pool can be quickly and efficiently narrowed to a manageable number of candidate germplasm for use in more focused and thorough studies to identify individual tolerance traits. This protocol subjects the plants to a rapidly induced severe water-stress in contrast to a more natural slowly-induced moderate continuous drought-stress that would be observed under field conditions. The type of response induced within the plant during these two types of water-stress events (rapid dehydration versus natural drought) is not identical9 which could lead to the exclusion of some materials from future trials. These two stress events are not without overlap though10 and this protocol should serve as an effective way to identify potentially drought-tolerant germplasm contained within a large collection. This protocol alone should not be considered sufficient to identify with a high degree of certainty germplasm that contains durable drought-tolerance under field water-stress conditions, but it represents a significant step forward in the rapid screening of leafy green vegetables for potential drought-tolerance traits.

<table border="0" cellpadding="0" cellspacing="0" width="791">
	<tbody>
		<tr height="54">
			<td height="54" style="height:54px;width:171px;">Name of Material/ equipment</td>
			<td style="width:135px;">Manufacturer</td>
			<td style="width:121px;">Catalog number</td>
			<td style="width:188px;">Comments</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;">plug tray 128</td>
			<td>T.O. Plastics 
			Hummert International</td>
			<td>11-8595-1</td>
			<td style="width:188px;">Any brand plug tray will work, but use the same style of trays for all trials.</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">lower tray (Display tray)</td>
			<td>T.O. Plastics 
			Hummert International</td>
			<td>11-3305-1</td>
			<td style="width:188px;"></td>
		</tr>
		<tr height="126">
			<td height="126" style="height:126px;width:171px;">plug/planting mix (Sunshine Mix #5)</td>
			<td>Sunshine 
			Hummert International</td>
			<td>10-0467-1</td>
			<td style="width:188px;">A different mix may need to be substituted if adapting this protocol to a different crop.&#160; Sunshine mix #4 was used in spinach trials.</td>
		</tr>
		<tr height="84">
			<td height="84" style="height:84px;">fertilizer (20-20-20)</td>
			<td>Jack&#39;s: Professional water-soluble fertilizer 
			Hummert International</td>
			<td>07-5915-1</td>
			<td style="width:188px;">Any fertilizer can be used, adjust type as needed for adapting this protocol to specific crop needs.</td>
		</tr>
	</tbody>
</table>

semi-high throughput screening for potential drought-tolerance in lettuce (lactuca sativa) germplasm collections

This protocol describes a method by which a large collection of the leafy green vegetable lettuce (Lactuca sativa L.) germplasm was screened for likely drought-tolerance traits. Fresh water availability for agricultural use is a growing concern across the United States as well as many regions of the world. Short-term drought events along with regulatory intervention in the regulation of water availability coupled with the looming threat of long-term climate shifts that may lead to reduced precipitation in many important agricultural regions has increased the need to hasten the development of crops adapted for improved water use efficiency in order to maintain or expand production in the coming years. This protocol is not meant as a step-by-step guide to identifying at either the physiological or molecular level drought-tolerance traits in lettuce, but rather is a method developed and refined through the screening of thousands of different lettuce varieties. The nature of this screen is based in part on the streamlined measurements focusing on only three water-stress indicators: leaf relative water content, wilt, and differential plant growth following drought-stress. The purpose of rapidly screening a large germplasm collection is to narrow the candidate pool to a point in which more intensive physiological, molecular, and genetic methods can be applied to identify specific drought-tolerant traits in either the lab or field. Candidates can also be directly incorporated into breeding programs as a source of drought-tolerance traits.

Bu protokol kuraklığa stres yanıtı halinde, istenilen deneysel özellik bir nüfusa ayırmak için büyük bir ekran yaparken, buna çok çeşitlidir olacak oluşturulan veriler muhtemel kuraklık toleranslı ve aradaki tüm noktalarda duyarlı stres. Şekil 1, bu protokol beklenen sonuçlara türünü temsil eden grafikler içeriyor. Üç farklı marul türlerine (marul (COS), marullarda ve Butterhead) temsili çeşidi sunulan veriler dahil edilmiştir. Sadece üç marul türleri temsil eden verile...

Watch this Scientific Journal Video about Semi-High Throughput Screening for Potential Drought-tolerance in Lettuce Lactuca sativa Germplasm Collections at JoVE.com

Semi-High Throughput Screening for Potential Drought-tolerance in Lettuce Lactuca sativa Germplasm Collections

This protocol was developed to screen a large germplasm collection of the leafy vegetable lettuce (Lactuca sativa L.) for drought-tolerance, in order to identify a small candidate pool of lettuce for use in physiological, molecular, and genetic studies to identify underlying drought-tolerance traits along with breeding programs.

Marul Potansiyel Kuraklığa tolerans Yarı-Yüksek Verim Tarama (<em&gt; Lactuca sativa</em&gt;) Germplazm Koleksiyonlar

This protocol describes a method by which a large collection of the leafy green vegetable lettuce (Lactuca sativa L.) germplasm was screened for likely ...

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8.9K Görüntüleme.  United States Department of Agriculture. This protocol was developed to screen a large germplasm collection of the leafy vegetable lettuce (Lactuca sativa L.) for drought-tolerance, in order to identify a small candidate pool of lettuce for use in physiological, molecular, and genetic studies to identify underlying drought-tolerance traits along with breeding programs.