Name: growing magnetotactic bacteria of the genus magnetospirillum: strains msr-1, amb-1 and ms-1
Uploaded: 2018-10-17T15:00:00
Description: Watch this Scientific Journal Video about Growing Magnetotactic Bacteria of the Genus Magnetospirillum: Strains MSR-1, AMB-1 and MS-1 at JoVE.com

我们感谢理查德 b. 弗兰克尔对山地车文化的帮助, 亚当. 希区柯克和朱慧在在麦克马斯特大学建立山地车文化, 以及为训练和访问电子显微设备 (麦克马斯特大学) 而进行的培训和使用。健康科学学院)。这项工作得到了加拿大自然科学和工程研究理事会 (NSERC) 和美国国家科学基金会的支持。

1. 安装 N2工位
注: 选择管的内径, 使其可以连接到最小泄漏的油箱, 使1毫升塑料注射器的气缸紧紧地装在管子中。图 1提供了完整的 N2气站示意图。
<ol><li>安全地安装一个 n2油箱靠近一个长凳上有足够的空间来设置 n2站 (长度约50厘米)。</li>
	<li>连接到水箱的一段足够长的管道, 以达到该站将建立的区域。如有必要, 在油?...

<ol>
	<li>Blakemore, R. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Magnetotactic+bacteria.">Magnetotactic bacteria.</a> Annual Reviews in Microbiology. 36 (1), 217-238 (1982).</li><li>Uebe, R., Sch&#252;ler, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Magnetosome+biogenesis+in+magnetotactic+bacteria.">Magnetosome biogenesis in magnetotactic bacteria.</a> Nature Reviews Microbiology. 14 (10), 621 (2016).</li><li>Faivre, D., Schuler, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Magnetotactic+bacteria+and+magnetosomes.">Magnetotactic bacteria and magnetosomes.</a> Chemical Reviews. 108 (11), 4875-4898 (2008).</li><li>Bazylinski, D. A., 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=Controlled+biomineralization+of+magnetite+(Fe3O4)+and+greigite+(Fe3S4)+in+a+magnetotactic+bacterium.">Controlled biomineralization of magnetite (Fe3O4) and greigite (Fe3S4) in a magnetotactic bacterium.</a> Applied and Environmental Microbiology. 61 (9), 3232-3239 (1995).</li><li>Lef&#232;vre, C. 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=Diversity+of+magneto-aerotactic+behaviors+and+oxygen+sensing+mechanisms+in+cultured+magnetotactic+bacteria.">Diversity of magneto-aerotactic behaviors and oxygen sensing mechanisms in cultured magnetotactic bacteria.</a> Biophysical Journal. 107 (2), 527-538 (2014).</li><li>Heyen, U., Sch&#252;ler, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Growth+and+magnetosome+formation+by+microaerophilic+Magnetospirillum+strains+in+an+oxygen-controlled+fermentor.">Growth and magnetosome formation by microaerophilic Magnetospirillum strains in an oxygen-controlled fermentor.</a> Applied Microbiology and Biotechnology. 61 (5-6), 536-544 (2003).</li><li>Blakemore, R. P., Maratea, D., Wolfe, R. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Isolation+and+pure+culture+of+a+freshwater+magnetic+spirillum+in+chemically+defined+medium.">Isolation and pure culture of a freshwater magnetic spirillum in chemically defined medium.</a> Journal of bacteriology. 140 (2), 720-729 (1979).</li><li>Oestreicher, Z., Lower, S. K., Lin, W., Lower, B. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Collection,+isolation+and+enrichment+of+naturally+occurring+magnetotactic+bacteria+from+the+environment.">Collection, isolation and enrichment of naturally occurring magnetotactic bacteria from the environment.</a> Journal of Visualized Experiments. (69), (2012).</li><li>P&#243;sfai, M., Lef&#232;vre, M., Trubitsyn, C., Bazylinski, D. A., Frankel, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Phylogenetic+significance+of+composition+and+crystal+morphology+of+magnetosome+minerals.">Phylogenetic significance of composition and crystal morphology of magnetosome minerals.</a> Frontiers in Microbiology. 4, 344 (2013).</li><li>Wolfe, R. S., Thauer, R. K., Pfennig, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+'capillary+racetrack'+method+for+isolation+of+magnetotactic+bacteria.">A 'capillary racetrack' method for isolation of magnetotactic bacteria.</a> FEMS Microbiology Ecology. 3 (1), 31-35 (1987).</li><li>Sch&#252;bbe, S., 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=Characterization+of+a+spontaneous+nonmagnetic+mutant+of+Magnetospirillum+gryphiswaldense+reveals+a+large+deletion+comprising+a+putative+magnetosome+island.">Characterization of a spontaneous nonmagnetic mutant of Magnetospirillum gryphiswaldense reveals a large deletion comprising a putative magnetosome island.</a> Journal of Bacteriology. 185 (19), 5779-5790 (2003).</li><li>Nadkarni, R., Barkley, S., Fradin, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+comparison+of+methods+to+measure+the+magnetic+moment+of+magnetotactic+bacteria+through+analysis+of+their+trajectories+in+external+magnetic+fields.">A comparison of methods to measure the magnetic moment of magnetotactic bacteria through analysis of their trajectories in external magnetic fields.</a> PloS One. 8 (12), e82064 (2013).</li><li>Waisbord, N., Lef&#232;vre, C. T., Bocquet, L., Ybert, C., Cottin-Bizonne, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Destabilization+of+a+flow+focused+suspension+of+magnetotactic+bacteria.">Destabilization of a flow focused suspension of magnetotactic bacteria.</a> Physical Review Fluids. 1 (5), 053203 (2016).</li><li>Komeili, A., Vali, H., Beveridge, T. J., Newman, D. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Magnetosome+vesicles+are+present+before+magnetite+formation,+and+MamA+is+required+for+their+activation.">Magnetosome vesicles are present before magnetite formation, and MamA is required for their activation.</a> Proceedings of the National Academy of Sciences of the United States of America. 101 (11), 3839-3844 (2004).</li><li>Scheffel, A., 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=An+acidic+protein+aligns+magnetosomes+along+a+filamentous+structure+in+magnetotactic+bacteria.">An acidic protein aligns magnetosomes along a filamentous structure in magnetotactic bacteria.</a> Nature. 440 (7080), 110 (2006).</li><li>Zhu, X., 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=Measuring+spectroscopy+and+magnetism+of+extracted+and+intracellular+magnetosomes+using+soft+X-ray+ptychography.">Measuring spectroscopy and magnetism of extracted and intracellular magnetosomes using soft X-ray ptychography.</a> Proceedings of the National Academy of Sciences of the United States of America. 113 (51), E8219-E8227 (2016).</li><li>Sch&#252;ler, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Molecular+analysis+of+a+subcellular+compartment:+the+magnetosome+membrane+in+Magnetospirillum+gryphiswaldense.">Molecular analysis of a subcellular compartment: the magnetosome membrane in Magnetospirillum gryphiswaldense.</a> Archives of Microbiology. 181 (1), 1-7 (2004).</li><li>Kolinko, I., 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=Biosynthesis+of+magnetic+nanostructures+in+a+foreign+organism+by+transfer+of+bacterial+magnetosome+gene+clusters.">Biosynthesis of magnetic nanostructures in a foreign organism by transfer of bacterial magnetosome gene clusters.</a> Nature Nanotechnology. 9 (3), 193 (2014).</li><li>Hergt, R., 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=Magnetic+properties+of+bacterial+magnetosomes+as+potential+diagnostic+and+therapeutic+tools.">Magnetic properties of bacterial magnetosomes as potential diagnostic and therapeutic tools.</a> Journal of Magnetism and Magnetic Materials. 293 (1), 80-86 (2005).</li><li>Lefevre, C. 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=Novel+magnetite-producing+magnetotactic+bacteria+belonging+to+the+Gammaproteobacteria.">Novel magnetite-producing magnetotactic bacteria belonging to the Gammaproteobacteria.</a> The ISME Journal. 6 (2), 440 (2012).</li><li>Williams, T. J., Lef&#232;vre, C. T., Zhao, W., Beveridge, T. J., Bazylinski, D. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Magnetospira+thiophila+gen.+nov.,+sp.+nov.,+a+marine+magnetotactic+bacterium+that+represents+a+novel+lineage+within+the+Rhodospirillaceae+(Alphaproteobacteria).">Magnetospira thiophila gen. nov., sp. nov., a marine magnetotactic bacterium that represents a novel lineage within the Rhodospirillaceae (Alphaproteobacteria).</a> International Journal of Systematic and Evolutionary Microbiology. 62 (10), 2443-2450 (2012).</li><li>Zhu, K., 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=Isolation+and+characterization+of+a+marine+magnetotactic+spirillum+axenic+culture+QH-2+from+an+intertidal+zone+of+the+China+Sea.">Isolation and characterization of a marine magnetotactic spirillum axenic culture QH-2 from an intertidal zone of the China Sea.</a> Research in Microbiology. 161 (4), 276-283 (2010).</li></ol>

山地车的特定 O2浓度要求使它们在实验室中生长不平凡。液体介质协议的一个关键步骤是从培养基中初始去除所有 o2 , 以通过在接种前添加一定体积的 o2来控制最终浓度。结果表明, MSR-1 生长在几乎完全有氧条件下, 然而, 细胞的磁性大大降低。同一研究结果表明, AMB-1 和 MS-1 在完全有氧条件下不生长6。对于 MSR-1 半固态培养, 所有 O2首先通过半胱?...

趋磁细菌 (MTB) 代表了广泛的革兰氏阴性原核生物在淡水和海洋水生生境中无处不在1。这些细菌共有能力产生磁性晶体由磁铁矿 (fe3O4) 或硫铁矿 (fe3S4), 这是在大多数情况下组装成链内的细胞。这个特定的结构主题是由于存在的几个特定的蛋白质作用于细菌的细胞质和周围的每个晶体2的脂膜。每个单独的晶体及其周围的膜泡被称为构建, 大小从大约30到 50 nm 在螺物种3。由于 magnetosomes 的链排列, 这些细菌具有永久的磁偶极矩, 使其与外部应用的磁场被动对齐。因此, 这些细菌积极地沿着磁场线游动, 作为自推进的微罗盘可能更有效地找到最有利的条件 (e., O2浓度) 增长。
山地车的一个有趣的特性是它们能够调节构建晶体的化学和晶体学。大多数菌株产生相对高纯度的磁铁或硫铁矿的晶体, 虽然一些 biomineralize 两个矿物4。在所有情况下, 细菌都能够精确地控制其单个磁性域晶体的大小和形状。这就解释了为什么需要进行大量的研究来更好地了解山地车如何执行这个生物矿化过程。了解这一过程可能使研究人员能够为许多商业和医疗应用量身定制磁性纳米晶体。
对山地车进行广泛研究的一个重大障碍是在实验室中种植它们的困难。大多数物种, 包括在这项工作中使用的菌株, 是好气微需氧当生长与 O2作为一个终端电子受体。这就解释了为什么这些细菌最常在好氧和缺氧条件 (好氧-缺氧界面, OAI) 之间的过渡区发现。这清楚地表明, 山地车具有精确的 O2浓度要求, 在为这些生物体设计生长培养基时, 显然需要考虑这些需求。此外, 山地车的巨大现有多样性意味着不同菌株将需要不同类型的化学梯度和养分来实现最佳生长。
在这项工作中, 我们描述了生长三个最广泛研究的山地车的方法:螺 magneticum (应变 AMB-1), m magnetotacticum (MS-1) 和m 螺(MSR-1)。这些物种复数属于Proteobacteria门的Alphaproteobacteria类, 在形态学上是螺旋状的, 在细胞的每个末端都具有极鞭毛。我们提供了在液体和 O2浓度梯度半固态介质中生长应变 MSR-1 的协议, 基于以前发布的培养基配方5,6。我们还介绍了改良磁性幽门生长培养基 (MGSM)7中生长菌株 AMB-1 和 MS-1 的详细协议。

<table border="0" cellpadding="0" cellspacing="0" style="width:1029px;" width="1028">
	<tbody>
		<tr height="22">
			<td height="22" style="height:23px;width:307px;">AMB-1</td>
			<td style="width:437px;">American Type Culture Collection (ATCC)</td>
			<td style="width:119px;">ATCC 700264</td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:23px;width:307px;">MS-1</td>
			<td>ATCC</td>
			<td>ATCC 31632&#160;</td>
			<td></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:23px;width:307px;">MSR-1</td>
			<td>Deutsche Sammlung von Mikroorganismen und Zellkulturen (DSMZ)</td>
			<td>DSM 6361</td>
			<td></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:23px;width:307px;">Ferric citrate</td>
			<td>Sigma-Aldrich</td>
			<td>F3388-250G</td>
			<td></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:23px;width:307px;">Trace mineral supplement</td>
			<td>ATCC</td>
			<td>MD-TMS</td>
			<td></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:23px;width:307px;">KH2PO4</td>
			<td>EMD</td>
			<td>PX1565-1</td>
			<td></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:23px;width:307px;">MgSO4.7 H2O</td>
			<td>EMD</td>
			<td>MX0070-1</td>
			<td></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:23px;width:307px;">HEPES</td>
			<td>BioShop Canada Inc</td>
			<td>HEP001.250</td>
			<td></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:23px;width:307px;">NaNO3</td>
			<td>Sigma-Aldrich</td>
			<td>S5506-250G</td>
			<td></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:23px;width:307px;">Yeast extract</td>
			<td>Fischer scientific</td>
			<td>DF210929</td>
			<td></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:23px;">Peptone</td>
			<td>Fischer scientific</td>
			<td>DF0436-17-5</td>
			<td></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:23px;width:307px;">Potassium L-lactate solution (60%)</td>
			<td>Sigma-Aldrich</td>
			<td>60389-250ML-F</td>
			<td></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:23px;width:307px;">D-(-)-Quinic acid</td>
			<td>Sigma-Aldrich</td>
			<td>138622</td>
			<td></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:23px;width:307px;">FeCl3.6H2O</td>
			<td>Fischer scientific</td>
			<td>I88-100</td>
			<td></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:23px;width:307px;">Vitamin supplement</td>
			<td>ATCC</td>
			<td>MD-VS</td>
			<td></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:23px;width:307px;">Sodium succinate hexahydrate</td>
			<td>Fischer scientific</td>
			<td>S413-500</td>
			<td></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:23px;width:307px;">Sodium L-tartrate dibasic dihydrate</td>
			<td>Sigma-Aldrich</td>
			<td>228729-100G</td>
			<td></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:23px;width:307px;">Sodium acetate trihydrate</td>
			<td>EMD</td>
			<td>SX0255-1</td>
			<td></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:23px;width:307px;">Resazurin</td>
			<td>Difco</td>
			<td>0704-13</td>
			<td></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:23px;width:307px;">Ascorbic acid</td>
			<td>Sigma-Aldrich</td>
			<td>A4544-25G</td>
			<td></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:23px;width:307px;">K2HPO4</td>
			<td>Caledon</td>
			<td>6620-1-65</td>
			<td></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:23px;">FeCl2 .4H2O</td>
			<td>Sigma-Aldrich</td>
			<td>44939-250G</td>
			<td></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:23px;width:307px;">Sodium bicarbonate</td>
			<td>EMD</td>
			<td>SX0320-1</td>
			<td></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:23px;width:307px;">NaCl</td>
			<td>Caledon</td>
			<td>7560-1</td>
			<td></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:23px;">NH4Cl</td>
			<td>EMD</td>
			<td>1011450500</td>
			<td></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:23px;">CaCl2.2 H2O</td>
			<td>EMD</td>
			<td>1023820500</td>
			<td></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:23px;width:307px;">Agar A</td>
			<td>Bio Basic Canada Inc</td>
			<td>FB0010</td>
			<td></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:23px;">L-cysteine.HCl.H2O</td>
			<td>Sigma-Aldrich</td>
			<td>C7880-100G</td>
			<td></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:23px;width:307px;">1.0 mL syringes</td>
			<td>Fischer scientific</td>
			<td>B309659</td>
			<td></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:23px;width:307px;">25G&#160; x 1 needles</td>
			<td>BD</td>
			<td>305125</td>
			<td></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:23px;width:307px;">125 mL serum bottles</td>
			<td>Wheaton</td>
			<td>223748</td>
			<td></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:23px;width:307px;">20 mm aluminum seals</td>
			<td>Wheaton</td>
			<td>224223-01</td>
			<td></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:23px;">20mm E-Z Crimper</td>
			<td>Wheaton</td>
			<td>W225303</td>
			<td></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:23px;width:307px;">Butyl-rubber stoppers</td>
			<td>Bellco Glass, Inc.</td>
			<td>2048-11800</td>
			<td></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:23px;width:307px;">Hungate tubes</td>
			<td>Chemglass (VWR)</td>
			<td>CLS-4208-01</td>
			<td></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:23px;width:307px;">Septum stopper, 13mm, Hungate</td>
			<td>Bellco Glass, Inc.</td>
			<td>2047-11600</td>
			<td></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:23px;width:307px;">Glass culture Tubes</td>
			<td>Corning (VWR)</td>
			<td>9826-16X</td>
			<td></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:23px;">Hydrochloric acid 36.5-38%, BioReagent</td>
			<td>Sigma-Aldrich</td>
			<td>H1758-100ML</td>
			<td>11.6 - 12 N</td>
		</tr>
	</tbody>
</table>

growing magnetotactic bacteria of the genus magnetospirillum: strains msr-1, amb-1 and ms-1

趋磁细菌是革兰氏阴性、运动性的, 主要水生原核生物在淡水和海洋生境中无处不在。它们的特点是他们的能力 biomineralize magnetosomes, 这是磁性纳米晶体的磁铁矿 (fe3O4) 或硫铁矿 (fe3S4) 包围的脂质双层膜, 在其细胞质。对于大多数已知的趋磁细菌, magnetosomes 被组装在细胞质内的链中, 从而赋予细胞永久的磁偶极矩, 并使它们与外部磁场被动对齐。由于这些特殊特性, 趋磁细菌对商业和医疗应用具有很大的潜力。然而, 大多数物种都是微需氧的, 有特定的 O2浓度要求, 使它们比其他许多细菌 (如大肠杆菌) 更难生长。在这里, 我们提出了生长在趋磁细菌的三种最广泛研究的菌株的详细的协议, 所有属于螺属。这些方法可以精确控制对细菌可用的 O2浓度, 以确保它们正常生长并合成 magnetosomes。使用这些程序进行进一步研究的趋磁细菌生长不需要实验家成为微生物学专家。本文中介绍的一般方法也可用于分离和培养其他趋磁细菌, 虽然生长培养基的化学成分可能需要修改。

成功制备培养基可按以下方式进行评估。在过程结束时, 应获得明确的解决方案 (不含任何沉淀) (这对于液体介质和 O2梯度半固态介质都是如此)。在接种前显示 MSR-1 液体培养基的预期方面的图片见图 2a。一个成功的 O2浓度梯度半固体介质是通过形成一个 OAI 后几个小时的信号, 表明由粉红色的介质在管的顶部的存在 (<strong class="...

Watch this Scientific Journal Video about Growing Magnetotactic Bacteria of the Genus Magnetospirillum: Strains MSR-1, AMB-1 and MS-1 at JoVE.com

Growing Magnetotactic Bacteria of the Genus Magnetospirillum: Strains MSR-1, AMB-1 and MS-1

螺属的生长趋磁菌: 菌株 MSR-1、AMB-1 和 MS-1

我们提出了在两种不同类型的生长培养基中生长几种螺菌株的程序。螺螺菌株 MSR-1 在液体和 O2浓度梯度半固态介质中生长, 而m magneticum应变 AMB-1 和m magnetotacticum应变 MS-1 在液体培养基中生长。

Magnetotactic bacteria are Gram-negative, motile, mainly aquatic prokaryotes ubiquitous in freshwater and marine habitats. They are characterized by their ...

这种方法允许种植三种不同种类的磁体细菌，称为MSR-1、AMB-1和MS-1。所有淡水物种，全部来自磁磷属。对于涉及这些生物体的任何可重复研究来说，这是重要的第一步。磁力细菌对氧气的要求非常特殊，这也是他们开发磁力导航系统的原因。因此，控制他们可用的氧气量是非常重要的。所有这些协议都针对一些特定的淡水磁螺旋菌株进行了优化。其细节可以优化，以适应其他磁体细菌的氧气和营养需求。在长凳附近安全安装氮气罐，该长凳上有足够的空间来设置氮气站。将一根管子连接到储罐，该油管足够长，可以到达空间站的建造区域。如有必要，在油箱输出时涂抹特氟隆胶带，以避免任何泄漏。要建造一个能够同时冒泡五瓶介质的站，切开四块长约五厘米的管子。将管子与三个 3 向 T 形塑料配件组装成一条线。通过额外的 T 形接头将此生产线的一端连接到氮气罐输出端的油管。在另一端添加 90 度弯头接头。使用胶带将结构连接到水平金属棒上，该金属棒位于长凳上方约 30 厘米处。将五条长度约 20 厘米的油管连接到已安装配件的三个输出。从五个一毫升塑料注射器中拆下活塞，并切割这些注射器的较大端，只保留分级部分。用棉花填充这些注射器，注意不要包装太紧。将注射器插入 20 厘米长的垂直管片中。打开气体，并使用肥皂水确保氮气流动时没有泄漏。取下五根 25 针的帽，并将这些针头插入 10 厘米的薄管中。将准备好的针头连接到氮气站的注射器上。确保氮气流经所有五条线路，并保持站内待命。通过将 0.245 克柠檬酸盐加入 100 毫升蒸馏去压水，准备 10 毫摩尔柠檬酸化液溶液。加热和搅拌溶解，直到获得橙色透明溶液。在121摄氏度下使用至少15分钟的暴露标准周期进行灭菌，然后在黑暗中将灭菌库存溶液储存在室温下。要为 MSR-1 准备五瓶液体生长介质，请将文本协议中列出的化学物质添加到含有 300 毫升蒸馏去水杯的烧杯中。在使用灭菌技术下，必须在Bunsen燃烧器的火焰下添加无菌柠檬酸盐和矿物溶液。无菌条件对于避免其他细菌物种对培养物的污染至关重要。添加所有化学品后，使用一种氢氧化钠溶液将pH调至7.0。将新鲜准备好的介质放入 125 毫升血清瓶中，在每个瓶子中倒入 60 毫升的介质。将氮气泡入介质 30 分钟，使用连接到氮站的小管清除溶解氧。在每个瓶子的顶部放置一个丁基橡胶塞，留下一个小开口，让多余的气体离开瓶子。30 分钟后，用准备好的塞子和铝密封密封每个瓶子。铝密封确保瓶子在协议的其余部分保持密封。断开针头和细管与氮气站的连接，用干净的针头更换。调节氮气罐的阀门，使温和、连续的气体流出油箱。现在，通过橡胶塞将连接到氮气罐的一根针头插入一瓶介质中。立即将另一根干净的针头插入同一瓶。对其他瓶子重复此步骤，让氮气流动约 30 分钟，用氮气代替瓶子中的空气。30 分钟后，通过取出相应的针头，将一瓶从氮气站断开。等待几秒钟，直到介质瓶中的压力减小到大气压力，然后取出第二根针。对所有剩余瓶子重复此步骤。要为 MSR-1 准备 120 毫升的半固体生长介质，请用铝箔盖住烧杯，并像文本协议中所述那样对解决方案进行解处理。就在自动处理器循环结束之前，准备一个新的 4%半胱氨酸溶液，并使用五摩尔氢氧化钠溶液将 pH 调整为 7.0，如文本协议中所述。高压灭式后，让介质冷却至50至60摄氏度，并将烧杯放在Bunsen燃烧器的火焰下。取出铝箔，在轻轻搅拌的同时，使用无菌技术快速添加文本协议中列出的化学物质。加入1.2毫升过滤器灭菌半胱氨酸溶液。添加所有化学品后，将加热介质转移到 16 毫升无菌螺帽 Hungate 管中。将12毫升的介质转移到每个管中，并密封管。要接种液体介质中的MSR-1、AMB-1和MS-1菌株，在塞子上涂乙醇并通过Bunsen燃烧器的火焰传递，点燃氧气和新鲜介质瓶的顶部。使用无菌注射器和针头，从氧气瓶中提取一毫升氧气，然后转移到新鲜的中瓶中。如果使用另一种培养在玻璃瓶中生长的培养剂，则点燃两个瓶子。然后将一毫升的较旧培养剂接种到新鲜培养中。在32摄氏度下孵育该培养，并在四至七天后接种到新鲜培养中。要接种氧气梯度、半固体介质中的 MSR-1，请验证新鲜介质管是否显示定义明确的 OAI，该管由粉红色到无色界面实现，该接口距离介质表面大约一到三厘米。如果注射液来自另一种氧气浓度梯度半固体培养物，则通过吸入培养物的50微升来收获细菌，并放置一个无菌移液器尖端，放在细菌形成的带子上。在OAI以新鲜介质中缓慢接种这些细菌，避免干扰界面。密封管子，让细菌生长在25摄氏度到30摄氏度之间。为了确保细菌既具有磁性，又具有模态性，请将一滴培养力放在显微镜盖上。翻转盖滑，并将其安装在放在玻璃滑梯上的 O 环上。确保跌落不接触底部幻灯片。将悬挂的落液放在显微镜下，并聚焦在液滴的边缘。将磁铁的南极靠近该边缘，观察细菌向边缘游去。翻转磁铁，让他们游向相反的方向。这里展示的是在接种前和细菌生长后，液体介质的瓶子。第二瓶的浊度表明细菌正在生长。这两次加速电影显示了模态和磁磁体细菌培养的悬挂滴实验的预期结果。当施加磁场时，细菌最初游向液滴的边缘。当磁体的磁场方向反转时，细菌游离施加磁场的边缘。这里显示的是在接种前和细菌生长后半固体介质管。不出所料，一带细菌在氧化-厌氧界面形成，随着氧气的消耗和界面向上移动而随时间而迁移。这里展示的是磁尖子的代表性电子显微图。旗子由黑色箭头指示，而红色箭头表示磁体。在尝试此过程时，重要的是要记住，磁体细菌有非常具体的氧气水平要求，以便正确生长和产生磁体体。该协议演示如何控制和监测生长介质中的氧含量，以实现这些生物体所首选的微观条件。按照这种方法，可以获得高浓度的健康和高磁性细胞。这样，可以进行需要大量细菌的实验，如磁体提取、基因组和基因研究，或通过显微镜观察集体运动。在实验室中生长磁体细菌的可能性确实让科学家能够系统地探索这些生物体的迷人生化和物理特性。

Research

JoVE Journal

Biology

Growing Magnetotactic Bacteria of the Genus Magnetospirillum: Strains MSR-1, AMB-1 and MS-1

Purification of the M. magneticum Strain AMB-1 Magnetosome Associated Protein MamA&Delta;41

Purification of the M. magneticum Strain AMB-1 Magnetosome Associated Protein MamAÃƒÅ½Ã¢â‚¬Â41

纯化 M。 magneticum应变AMB - 1磁小体相关蛋白MamAΔ41

Magnetotactic bacteria comprise a diverse group of aquatic microorganisms that are able to orientate themselves along geomagnetic fields. This behavior is ...

科研

生物学

High-throughput Screening and Biosensing with Fluorescent C. elegans Strains

High-throughput Screening and Biosensing with Fluorescent C. elegans Strains

高通量筛选和生物传感与荧光 C。线虫菌株

High-throughput screening (HTS) is a powerful approach for identifying chemical modulators of biological processes.  However, many compounds identified in ...

The Green Monster Process for the Generation of Yeast Strains Carrying Multiple Gene Deletions

绿色怪物承载多基因缺失的酵母菌株的产生过程

Phenotypes  for a gene deletion are often revealed only when the mutation is tested in a  particular genetic background or environmental condition1,2. ...

Collection, Isolation and Enrichment of Naturally Occurring Magnetotactic Bacteria from the Environment

从环境中自然产生的趋磁细菌的采集，分离和富集

Magnetotactic bacteria (MTB) are aquatic microorganisms that were first notably described in 19751 from sediment samples collected in salt marshes of ...

Monitoring Intraspecies Competition in a Bacterial Cell Population by Cocultivation of Fluorescently Labelled Strains

通过共生荧光标记菌株监测细菌细胞群中的物种内部竞争

Many microorganisms such as bacteria proliferate extremely fast and the populations may reach high cell densities. Small fractions of cells in a ...

Measuring the Effects of Bacteria on C. Elegans Behavior Using an Egg Retention Assay

Measuring the Effects of Bacteria on C. Elegans Behavior Using an Egg Retention Assay

测量细菌的影响<em&gt; C。线虫</em&gt;行为使用一个鸡蛋保留含量

C. elegans egg-laying behavior is  affected by environmental cues such as osmolarity1 and  vibration2. In the total absence of food C. elegans also cease ...

Measurement of mRNA Decay Rates in Saccharomyces cerevisiae Using rpb1-1 Strains

Measurement of mRNA Decay Rates in Saccharomyces cerevisiae Using rpb1-1 Strains

mRNA的衰变率的测量<em&gt;酿酒酵母</em&gt;使用<em&gt; rpb1-1</em&gt;株

mRNA steady state levels vary depending on environmental conditions. Regulation of the steady state accumulation levels of an mRNA ensures that the ...

Using Digital Image Correlation to Characterize Local Strains on Vascular Tissue Specimens

利用数字图像相关表征的血管组织标本本地菌株

Characterization of the mechanical behavior of biological and engineered soft tissues is a central component of fundamental biomedical research and ...

Colorimetric Detection of Bacteria Using Litmus Test

细菌利用试金石比色法检测

There are increasing demands for simple but still effective methods that can be used to detect specific pathogens for point-of-care or field applications. ...

Light Sheet Fluorescence Microscopy of Plant Roots Growing on the Surface of a Gel

植物根系的轻质板材荧光显微镜生长在一个凝胶表面

One of the key questions in understanding plant development is how single cells behave in a larger context of the tissue. Therefore, it requires the ...