Name: automated microbial cultivation and adaptive evolution using microbial microdroplet culture system (mmc)
Uploaded: 2022-02-18T12:30:00
Description: Watch this Scientific Journal Video about Automated Microbial Cultivation and Adaptive Evolution using Microbial Microdroplet Culture System MMC at JoVE.com

本研究得到了国家重点研发计划（2018YFA0901500）、国家自然科学基金国家重点仪器设备专项（21627812）和清华大学自主研发计划（20161080108）的支持。我们还感谢Julia A. Vorholt教授（苏黎世联邦理工学院生物学系微生物研究所，苏黎世联邦理工学院，苏黎世8093，瑞士）提供甲醇必需 的大肠杆菌 菌株2.2版（MeSV2.2）。

1. 仪器和软件安装
<ol><li>选择清洁无菌环境（如洁净工作台）作为MMC的专用永久空间。在空间中稳定地安装MMC。 注意：使MMC远离强电场，磁场和强热辐射源的干扰。避免剧烈振动影响光学检测元件。向MMC提供AC220 V，50 HZ的电源。有关 MMC 的详细信息，请参阅 材料表 和 MMC16 的网站。</li>
	<li>从 MMC.zip 文件安装操作软件 注意：请与作者联系以获取 MMC...

<ol>
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该协议介绍了如何使用微生物滴培养系统（MMC）进行自动微生物培养和长期适应性进化。MMC是一种小型化，自动化和高通量微生物培养系统。与传统的微生物高通量培养方法和仪器相比，MMC具有劳动力和试剂消耗少、操作简单、在线检测（OD和荧光）、高时间分辨率数据采集、超并行化等诸多优点。MMC还具有一些与常规液滴微流体技术不同的特殊优势，后者通常使用pL和nL液滴。以前报道的大多数?...

微生物培养是微生物科学研究和工业应用的重要基础，广泛应用于微生物的分离、鉴定、重建、筛选和进化1，2，3。传统的微生物培养方法主要采用试管、摇瓶、实心板作为培养容器，结合振荡培养箱、分光光度计、酶标仪等设备进行微生物培养、检测和筛选。但是，这些方法存在许多问题，例如操作繁琐，通量低，效率低，消耗大量劳动力和试剂。近年来开发的高通量培养方法主要基于微孔板。但微孔板溶解氧水平低，混合性能差，蒸发和热效应严重，往往导致微生物生长状态和实验平行化不良4，5，6，7;另一方面，它需要配备昂贵的设备，如液体处理工作站和酶标仪，以实现自动化培养和过程检测8，9。
作为微流控技术的一个重要分支，液滴微流体近年来在传统的连续流微流体系统的基础上发展起来。它是一种离散流动微流体技术，它使用两个不混溶的液相（通常是油 - 水）来产生分散的微液滴并在其上运行10。由于微液滴具有体积小、比表面积大、内部传质率高、区室化不造成交叉污染等特点，以及液滴可控性强、通量高的优点，因此应用液滴微流控技术应用于微生物高通量培养、筛选和进化等已有多种研究11.然而，仍有一系列关键问题需要使液滴微流控技术普及和广泛应用。首先，液滴微流体的操作繁琐复杂，对操作人员的技术要求很高。其次，液滴微流控技术结合了光学、机械和电气元件，需要与生物技术应用场景相关联。如果没有多学科协作，单个实验室或团队就很难建立高效的液滴微流体控制系统。第三，由于微液滴体积小（从皮升（pL）到微升（μL）），对于一些基本的微生物操作，如亚培养、分选、取样，实现液滴的精确自动化控制和实时在线检测需要很大的难度，并且难以构建一体化设备系统12。
为了解决上述问题，基于液滴微流控技术成功开发了一种全自动微生物微滴培养系统（MMC）。MMC由四个功能模块组成：液滴识别模块、液滴光谱检测模块、微流控芯片模块和采样模块。通过所有模块的系统集成和控制，准确建立包括液滴生成、培养、测量（光密度（OD）和荧光）、分裂、融合、分选在内的自动化操作系统，实现微生物液滴培养过程中所需的接种、培养、监测、分培养、分选、采样等功能的集成。MMC可容纳多达200个2-3μL体积的重复液滴培养单元，相当于200个摇瓶培养单元。微滴培养系统可以满足微生物生长过程中无污染、溶解氧、混合、质能交换的要求，通过生长曲线测量、自适应进化、单因子多级分析、代谢物研究与分析（基于荧光检测）等多种集成功能，满足微生物研究的各种需求13，14。
在这里，该协议详细介绍了如何使用MMC进行自动化和微生物培养以及适应性进化（图1）。以野生型大肠杆菌MG1655为例，以生长曲线测量和甲醇必需的大肠杆菌菌株MesV2.215为例，证明了MMC的适应性进化。开发了MMC的操作软件，使操作非常简单明了。在整个过程中，用户需要准备初始细菌溶液，设置MMC的条件，然后将细菌溶液和相关试剂注入MMC。随后，MMC将自动执行液滴生成，识别和编号，培养和自适应进化等操作。它还将以高时间分辨率对液滴进行在线检测（OD和荧光），并在软件中显示相关数据（可以导出）。操作员可以根据结果随时停止培养过程，并提取目标液滴以进行后续实验。MMC操作简便，消耗的劳动力和试剂少，实验通量相对较高，数据并行度好，与常规培养方法相比具有显著优势。它为研究人员进行相关的微生物研究提供了一个低成本，操作友好且强大的实验平台。

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>0.22 &#956;m PVDF filter membrane</td>
			<td>Merck Millipore Ltd.</td>
			<td>SLGPR33RB</td>
			<td>Sterilize the MMC oil</td>
		</tr>
		<tr>
			<td>4 &#176;C refrigerator</td>
			<td>Haier</td>
			<td>BCD-289BSW</td>
			<td>For reagent storage</td>
		</tr>
		<tr>
			<td>Agar</td>
			<td>Becton, Dickinson and Company</td>
			<td>214010</td>
			<td>For solid plate preparation</td>
		</tr>
		<tr>
			<td>CaCl2&#183;2H2O</td>
			<td>Sinopharm Chemical Reagent Beijing Co., Ltd.</td>
			<td>20011160</td>
			<td>Component of the special medium for MeSV2.2.</td>
		</tr>
		<tr>
			<td>Clean bench</td>
			<td>Beijing Donglian Har Instrument Manufacture Co., Ltd.</td>
			<td>DL-CJ-INDII</td>
			<td>For aseptic operation and UV sterilization</td>
		</tr>
		<tr>
			<td>CoCl2&#183;6H2O</td>
			<td>Sinopharm Chemical Reagent Beijing Co., Ltd.</td>
			<td>10007216</td>
			<td>Component of the special medium for MeSV2.2.</td>
		</tr>
		<tr>
			<td>Computer</td>
			<td>Lenovo</td>
			<td>E450</td>
			<td>Software installation and MMC control</td>
		</tr>
		<tr>
			<td>Constant temperature incubator</td>
			<td>Shanghai qixin scientific instrument co., LTD</td>
			<td>LRH 250</td>
			<td>For the microbial cultivation using solid medium</td>
		</tr>
		<tr>
			<td>CuSO4&#183;5H2O</td>
			<td>Sinopharm Chemical Reagent Beijing Co., Ltd.</td>
			<td>10008218</td>
			<td>Component of the special medium for MeSV2.2.</td>
		</tr>
		<tr>
			<td>Electronic balance</td>
			<td>OHAUS</td>
			<td>AR 3130</td>
			<td>For reagent weighing</td>
		</tr>
		<tr>
			<td>EP tube</td>
			<td>Thermo Fisher</td>
			<td>1.5 mL</td>
			<td>For droplet collection</td>
		</tr>
		<tr>
			<td>FeCl3&#183;6H2O</td>
			<td>Sinopharm Chemical Reagent Beijing Co., Ltd.</td>
			<td>10011928</td>
			<td>Component of the special medium for MeSV2.2.</td>
		</tr>
		<tr>
			<td>Freezing Tube</td>
			<td>Thermo Fisher</td>
			<td>2.0 mL</td>
			<td>For strain preservation</td>
		</tr>
		<tr>
			<td>Gluconate</td>
			<td>Sigma-Aldrich</td>
			<td>S2054</td>
			<td>Component of the special medium for MeSV2.2.</td>
		</tr>
		<tr>
			<td>Glycerol</td>
			<td>GENERAL-REAGENT</td>
			<td>G66258A</td>
			<td>For strain preservation</td>
		</tr>
		<tr>
			<td>High-Pressure Steam Sterilization Pot</td>
			<td>SANYO Electric</td>
			<td>MLS3020</td>
			<td>For autoclaved sterilization</td>
		</tr>
		<tr>
			<td>isopropyl-&#946;-d-thiogalactopyranoside (IPTG)</td>
			<td>Biotopped</td>
			<td>420322</td>
			<td>Component of the special medium for MeSV2.2.</td>
		</tr>
		<tr>
			<td>Kanamycin sulfate</td>
			<td>Solarbio</td>
			<td>K8020</td>
			<td>Component of the special medium for MeSV2.2.</td>
		</tr>
		<tr>
			<td>KH2PO4</td>
			<td>MACKLIN</td>
			<td>P815661</td>
			<td>Component of the special medium for MeSV2.2.</td>
		</tr>
		<tr>
			<td>Methanol</td>
			<td>MACKLIN</td>
			<td>M813895</td>
			<td>Component of the special medium for MeSV2.2.</td>
		</tr>
		<tr>
			<td>MgSO4&#183;7H2O</td>
			<td>BIOBYING</td>
			<td>1305715</td>
			<td>Component of the special medium for MeSV2.2.</td>
		</tr>
		<tr>
			<td>Microbial Microdroplet Culture System (MMC)</td>
			<td>Luoyang TMAXTREE Biotechnology Co., Ltd.</td>
			<td>&#160;MMC-I</td>
			<td>Performing growth curve determination and adaptive evolution. Please refer to http://www.tmaxtree.com/en/index.php?v=news&#38;id=110</td>
		</tr>
		<tr>
			<td>Microfluidic chip</td>
			<td>Luoyang TMAXTREE Biotechnology Co., Ltd.</td>
			<td>MMC-ALE-OD</td>
			<td>For various droplet operations. Please refer to http://www.tmaxtree.com/en/</td>
		</tr>
		<tr>
			<td>MMC oil</td>
			<td>Luoyang TMAXTREE Biotechnology Co., Ltd.</td>
			<td>MMC-M/S-OD</td>
			<td>The oil phase for droplet microfluidics. Please refer to http://www.tmaxtree.com/en/</td>
		</tr>
		<tr>
			<td>MnCl2</td>
			<td>Sinopharm Chemical Reagent Beijing Co., Ltd.</td>
			<td>20026118</td>
			<td>Component of the special medium for MeSV2.2.</td>
		</tr>
		<tr>
			<td>NaCl</td>
			<td>GENERAL-REAGENT</td>
			<td>G81793J</td>
			<td>Component of the LB medium</td>
		</tr>
		<tr>
			<td>Na2HPO4&#183;12H2O</td>
			<td>GENERAL-REAGENT</td>
			<td>G10267B</td>
			<td>Component of the special medium for MeSV2.2.</td>
		</tr>
		<tr>
			<td>NH4Cl</td>
			<td>Sinopharm Chemical Reagent Beijing Co., Ltd.</td>
			<td>10001518</td>
			<td>Component of the special medium for MeSV2.2.</td>
		</tr>
		<tr>
			<td>Petri dish</td>
			<td>Corning Incorporated</td>
			<td>90 mm</td>
			<td>For the preparation of solid medium</td>
		</tr>
		<tr>
			<td>Pipette</td>
			<td>eppendorf</td>
			<td>2.5 &#956;L, 10 &#956;L, 100&#956;L, 1000&#956;L</td>
			<td>For liquid handling</td>
		</tr>
		<tr>
			<td>Quick connector A</td>
			<td>Luoyang TMAXTREE Biotechnology Co., Ltd.</td>
			<td>&#8212;</td>
			<td>For the connection of each joint. Please refer to http://www.tmaxtree.com/en/</td>
		</tr>
		<tr>
			<td>Reagent bottle</td>
			<td>Luoyang TMAXTREE Biotechnology Co., Ltd.</td>
			<td>MMC-PCB</td>
			<td>Sampling and storage of bacteria solution and reagents. Please refer to http://www.tmaxtree.com/en/</td>
		</tr>
		<tr>
			<td>Shake flask</td>
			<td>Union-Biotech</td>
			<td>50 mL</td>
			<td>For microbial cultivation</td>
		</tr>
		<tr>
			<td>Shaking incubator</td>
			<td>Shanghai Sukun Industrial Co., Ltd.</td>
			<td>SKY-210 2B</td>
			<td>For the microbial cultivation in shake flask</td>
		</tr>
		<tr>
			<td>Streptomycin sulfate</td>
			<td>Solarbio</td>
			<td>S8290</td>
			<td>Component of the special medium for MeSV2.2.</td>
		</tr>
		<tr>
			<td>Syringe</td>
			<td>JIANGSU ZHIYU MEDICAL INSTRUCTMENT CO., LTD</td>
			<td>10 mL</td>
			<td>Draw liquid and inject it into the reagent bottle</td>
		</tr>
		<tr>
			<td>Syringe needle</td>
			<td>OUBEL Hardware Store</td>
			<td>22G</td>
			<td>Inner diameter is 0.41 mm and outer diameter is 0.71 mm.</td>
		</tr>
		<tr>
			<td>Tryptone</td>
			<td>Oxoid Ltd.</td>
			<td>LP0042</td>
			<td>Component of the LB medium</td>
		</tr>
		<tr>
			<td>Ultra low temperature refrigerator</td>
			<td>SANYO Ultra-low</td>
			<td>MDF-U4086S</td>
			<td>For strain preservation (-80 &#176;C)</td>
		</tr>
		<tr>
			<td>UV&#8211;Vis spectrophotometer</td>
			<td>General Electric Company</td>
			<td>Ultrospec 3100 pro</td>
			<td>For the measurement of OD values</td>
		</tr>
		<tr>
			<td>Vitamin B1</td>
			<td>Solarbio</td>
			<td>SV8080</td>
			<td>Component of the special medium for MeSV2.2.</td>
		</tr>
		<tr>
			<td>Yeast extract</td>
			<td>Oxoid Ltd.</td>
			<td>LP0021</td>
			<td>Component of the LB medium</td>
		</tr>
		<tr>
			<td>ZnSO4&#183;7H2O</td>
			<td>Sinopharm Chemical Reagent Beijing Co., Ltd.</td>
			<td>10024018</td>
			<td>Component of the special medium for MeSV2.2.</td>
		</tr>
	</tbody>
</table>

automated microbial cultivation and adaptive evolution using microbial microdroplet culture system (mmc)

传统的微生物培养方法通常操作繁琐，通量低，效率低，消耗大量劳动力和试剂。此外，近年来开发的基于微孔板的高通量培养方法由于其溶解氧低，混合物不良以及严重的蒸发和热效应，微生物生长状况较差，实验平行化程度低。由于微液滴的诸多优点，如体积小、通量高、可控性强等，液滴基微流控技术可以克服这些问题，已应用于多种高通量微生物培养、筛选、进化的研究。然而，大多数先前的研究仍处于实验室建设和应用阶段。一些关键问题，如操作要求高，施工难度高，缺乏自动化集成技术，限制了液滴微流控技术在微生物研究中的广泛应用。在这里，基于液滴微流技术成功开发了一种自动化微生物微滴培养系统（MMC），实现了微生物液滴培养过程中所需的接种、培养、在线监测、亚培养、分选、取样等功能的集成。以野生型大肠杆菌MG1655和甲醇必需大肠杆菌菌株（MeSV2.2）为例，详细介绍了如何使用MMC进行自动化和相对高通量的微生物培养和适应性进化。该方法操作简便，消耗的人工和试剂少，实验通量高，数据并行度好，与常规培养方法相比具有很大的优势。它为科研人员开展相关微生物研究提供了一个低成本、操作友好、结果可靠的实验平台。

该协议使用 大肠杆菌 MG1655和MesV2.2菌株作为示例，以展示MMC中具有自动化和相对高通量策略的微生物培养和甲醇必需的适应性进化。生长曲线测量主要用于表征微生物培养。通过自动连续亚培养和在每个亚培养过程中加入高浓度甲醇作为选择压力进行适应性进化。通过每个次培养期间液滴最大OD值的变化趋势来估计是否实现了适应性进化。MMC的可调参数和精度参数如 表 2<...

Watch this Scientific Journal Video about Automated Microbial Cultivation and Adaptive Evolution using Microbial Microdroplet Culture System MMC at JoVE.com

Automated Microbial Cultivation and Adaptive Evolution using Microbial Microdroplet Culture System MMC

该协议描述了如何使用微生物滴培养系统（MMC）进行自动微生物培养和适应性进化。MMC可以自动连续地培养和分培养微生物，并在线监测其生长，具有相对较高的通量和良好的并行化性，减少了劳动力和试剂消耗。

使用微生物滴培养系统（MMC）的自动微生物培养和适应性进化

Conventional microbial cultivation methods usually have cumbersome operations, low throughput, low efficiency, and large consumption of labor and ...

这种方法可以帮助解决常规微生物培养方法的问题。它为微生物的自动化培养和适应性进化提供了低成本，操作友好且结果可靠的实验平台。该技术实现了操作自动化，并进一步降低了劳动力消耗。同时，它具有良好的培养性能，结果可靠，操作简单。该技术主要应用于微生物学研究领域，如生长曲线测量、自适应实验室进化、单因素多层次分析等。通过将内径为0.41毫米，外径为0.71毫米的注射器针头连接到快速接头A和试剂瓶上，开始实验的准备工作。然后将所有设备在121摄氏度下高压灭菌15分钟。要安装微流控芯片，请打开操作室的门并抬起光纤探头。将电场孔与针对齐后，轻轻地将芯片放在芯片基座上，然后将两根定位柱插入定位孔中，放下光纤探头。接下来，根据手稿中描述的位置编号，将芯片上的快速连接器A连接到微生物微滴培养系统或MMC的相应端口。完成后，关闭操作室的门。用培养基将培养的大肠杆菌MG1655悬浮液稀释至0.05至0.1的OD 600，以获得10毫升初始细菌溶液。要初始化 MMC，请单击“初始化”选项卡。当初始化界面出现时，将培养温度设置为37摄氏度，将光电信号值设置为0.6。初始化大约需要 20 分钟。之后，将灭菌的试剂瓶放在洁净工作台上并拧紧盖子。使用10毫升无菌注射器将三至五毫升MMC油从注射器针头注射到侧管中放入试剂瓶中。然后缓慢倾斜并旋转试剂瓶，使油完全渗入内壁。将五毫升初始细菌溶液注入瓶中后，通过注入五至七毫升MMC油来填充试剂瓶。拉出试剂瓶的独立快速接头A，将快速接头A插入瓶的快速接头B中，完成进样操作。完成后，打开操作室的门，将试剂瓶放入金属浴中。拉出芯片的C2连接器和试剂瓶的快速连接器A。将试剂瓶的侧管连接器连接到 C2 连接器，将顶管连接器连接到 O2 连接器。然后关闭手术室的门。要选择生长曲线测量功能，请单击参数设置界面中的生长曲线，输入数字为15。然后打开OD检测开关，将波长设置为600纳米。单击“开始”选项卡以开始生成液滴。该过程将需要 15 分钟才能完成。当主界面上出现提示消息的弹出窗口时，打开操作室的门取出试剂瓶并连接C2和O2连接器。关上门后，单击弹出窗口中的“确定”按钮，自动培养液滴并检测OD值。当增长曲线达到静止阶段时，单击“数据导出”按钮以导出 OD 数据。选择数据保存路径，并以CVS格式导出栽培期间记录的OD值 要绘制生长曲线，请使用Excel和Origin 9.0等绘图软件。如前所述，将所需体积的初始细菌溶液，新鲜培养基和MMC油注入单独的灭菌试剂瓶中，用于新鲜培养基中的初始细菌溶液。要选择自适应进化的功能，请单击软件中的ALE。在参数设置界面中，打开OD检测开关，然后设置稿件中描述的所有参数，点击“开始”选项卡，开始液滴生成。该过程大约需要25分钟。在每个子培养期间，观察液滴的最大OD值是否显着增加。如果发生增加并满足实验要求，请单击数据导出按钮以导出OD数据。要从MMC中提取目标液滴，请单击筛选选项卡以选择液滴提取功能。然后选择“收集”选项并单击目标液滴的数量。完成后，单击“确定”。等待提示消息的弹出窗口。然后将CF快速接头放入微量离心管中进行收集，然后单击“确定”。一到两分钟后，当软件界面会弹出一个新窗口提示消息时，将CF快速连接器插入回去，然后单击“确定”以使MMC继续运行。当下一个目标液滴到达液滴识别位点时，按照之前的规定收集它。使用2.5微升移液器取出液滴并将其置于90毫米实心板上，然后用侧面长度为3厘米的玻璃三角形涂层棒均匀地铺展液滴，然后在37摄氏度恒温培养箱中培养液滴72小时。之后，选择三到五个独立的细菌菌落，在50毫升的摇瓶中分别培养每个菌落，在摇动培养箱中以每分钟200转和37摄氏度培养48至72小时。孵育后，按照相关标准规定将培养的细菌溶液储存在甘油管中。代表性分析显示了50个液滴在整个适应性进化过程中的生长曲线。观察到甲醇必需大肠杆菌菌株或MeSV2.2显示出初始缓慢生长，随后快速生长。分别绘制了6号液滴在整个适应性进化过程中的生长曲线。第一代最大OD600值为0.37，在上一个次培养期增加到0.58，表明液滴6中的菌株实现了明显的适应性进化。此外，比较了液滴6菌株和初始菌株的生长曲线。液滴6菌株在固定期表现出比初始菌株更高的最大比生长速率和细胞浓度。最重要的是要保证仪器芯片与试剂瓶的连接正确，这是操作正常进行的液滴。该方法为研究人员提供了微生物培养的新思路，也为菌株的适应性进化提供了高效、低成本的平台。

Research

JoVE Journal

Bioengineering

Automated Microbial Cultivation and Adaptive Evolution using Microbial Microdroplet Culture System (MMC)

Microfluidic Picoliter Bioreactor for Microbial Single-cell Analysis: Fabrication, System Setup, and Operation

In this protocol the fabrication, experimental setup and basic operation of the recently introduced microfluidic picoliter bioreactor (PLBR) is described ...

科研

生物工程

Cultivation of Mammalian Cells Using a Single-use Pneumatic Bioreactor System

培养哺乳动物细胞使用一次性使用气动生物反应器系统

Recent advances in mammalian, insect, and stem cell cultivation and scale-up have created tremendous opportunities for new therapeutics and personalized ...

Electrotaxis Studies of Lung Cancer Cells using a Multichannel Dual-electric-field Microfluidic Chip

使用多通道双电场微流控芯片肺癌细胞Electrotaxis研究

The behavior of directional cell migration under a direct current electric-field (dcEF) is referred to as electrotaxis. The significant role of ...

A Novel Bioreactor for High Density Cultivation of Diverse Microbial Communities

一种新型生物反应器为不同的微生物群落的高密度栽培

A novel reactor design, coined a high density bioreactor (HDBR), is presented for the cultivation and study of high density microbial communities. Past ...

Design and Implementation of an Automated Illuminating, Culturing, and Sampling System for Microbial Optogenetic Applications

设计和自动照明，培养和取样系统实现微生物光遗传学的应用

Optogenetic systems utilize genetically-encoded proteins that change conformation in response to specific wavelengths of light to alter cellular ...

Assembly and Tracking of Microbial Community Development within a Microwell Array Platform

微孔阵列平台中微生物群落发展的组装与跟踪

The development of microbial communities depends on a combination of complex deterministic and stochastic factors that can dramatically alter the spatial ...

Prospecting Microbial Strains for Bioremediation and Probiotics Development for Metaorganism Research and Preservation

生物修复和益生菌发育的微生物菌株的勘探，用于生物有机物的研究与保存

Pollution affects all biomes. Marine environments have been particularly impacted, especially coral reefs, one of the most sensitive ecosystems on Earth. ...

Process Optimization using High Throughput Automated Micro-Bioreactors in Chinese Hamster Ovary Cell Cultivation

中国仓鼠卵巢细胞培养中高通量自动化微生物反应器工艺优化

Optimization of bioprocesses to increase the yield of desired products is of importance in the biopharmaceutical industry. This can be achieved by strain ...

Generating Controlled, Dynamic Chemical Landscapes to Study Microbial Behavior

生成受控动态化学景观，研究微生物行为

We demonstrate a method for the generation of controlled, dynamic chemical pulses&#8213;where localized chemoattractant becomes suddenly available at the ...

High-Throughput Identification of Resistance to Pseudomonas syringae pv. Tomato in Tomato using Seedling Flood Assay

High-Throughput Identification of Resistance to Pseudomonas syringae pv. Tomato in Tomato using Seedling Flood Assay

高通量识别对伪单鱼的电阻syringae pv。番茄中的番茄使用苗子洪水测定

Tomato is an agronomically important crop that can be infected by Pseudomonas syringae, a Gram-negative bacterium, resulting in bacterial speck disease. ...