Name: generation of fluorescent protein fusions in candida species
Uploaded: 2017-03-04T15:00:00
Description: Watch this Scientific Journal Video about Generation of Fluorescent Protein Fusions in Candida Species at JoVE.com

我们感谢N.院长提供原mCherry FP序列，M Gerami - 内贾德建设质粒，拉尔森B.技术援助和T.海泽尔这个项目的开发过程中有益的建议。 JB是由欧洲研究委员会高级奖340087（RAPLODAPT）的支持。显微及成像系统是由明尼苏达州儿科基金会大学和明尼苏达影像中心的大学提供。

1.切断模板质粒从大肠杆菌 <ol><li>生长含模板质粒过夜在10ml LB培养基（LB）+ 200毫克/升氨苄青霉素（AMP），在37℃振荡的大肠杆菌 。 </li><li>收获细胞通过在6000 xg离心离心2分钟。 </li><li>倾析液体，分离和纯化的DNA从大肠杆菌如Ausubel 等人以前描述通过标准方法的大肠杆菌细胞。 8。 </li><li>在50-100 ng / ml的工作浓度;重悬的DNA在Tris-EDTA（10毫...

<ol>
	<li>Bendel, C. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Colonization+and+epithelial+adhesion+in+the+pathogenesis+of+neonatal+candidiasis.">Colonization and epithelial adhesion in the pathogenesis of neonatal candidiasis.</a> Semin. Perinatol. 27 (5), 357-364 (2003).</li><li>Gerami-Nejad, M., Berman, J., Gale, C. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cassettes+for+PCR-mediated+construction+of+green,+yellow,+and+cyan+fluorescent+protein+fusions+in+Candida+albicans.">Cassettes for PCR-mediated construction of green, yellow, and cyan fluorescent protein fusions in Candida albicans.</a> Yeast. 18 (9), 859-864 (2001).</li><li>Gerami-Nejad, M., Dulmage, K., Berman, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Additional+cassettes+for+epitope+and+fluorescent+fusion+proteins+in+Candida+albicans.">Additional cassettes for epitope and fluorescent fusion proteins in Candida albicans.</a> Yeast. 26 (7), 399-406 (2009).</li><li>Gerami-Nejad, M., Forche, A., McClellan, M., Berman, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Analysis+of+protein+function+in+clinical+C.+albicans+isolates.">Analysis of protein function in clinical C. albicans isolates.</a> Yeast. 29 (8), 303-309 (2012).</li><li>Gerami-Nejad, M., Hausauer, D., McClellan, M., Berman, J., Gale, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cassettes+for+the+PCR-mediated+construction+of+regulatable+alleles+in+Candida+albicans.">Cassettes for the PCR-mediated construction of regulatable alleles in Candida albicans.</a> Yeast. 21 (5), 429-436 (2004).</li><li>Gonia, S., Larson, B., Gale, C. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=PCR-mediated+gene+modification+strategy+for+construction+of+fluorescent+protein+fusions+in+Candida+parapsilosis.">PCR-mediated gene modification strategy for construction of fluorescent protein fusions in Candida parapsilosis.</a> Yeast. 33 (2), 63-69 (2016).</li><li>Milne, S. W., Cheetham, J., Lloyd, D., Aves, S., Bates, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cassettes+for+PCR-+mediated+gene+tagging+in+Candida+albicans+utilizing+nourseothricin+resistance.">Cassettes for PCR- mediated gene tagging in Candida albicans utilizing nourseothricin resistance.</a> Yeast. 28 (12), 833-841 (2011).</li><li>Ausubel, F. 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=.">.</a> Current Protocols in Molecular Biology. , (1995).</li><li>Wilson, R. B., Davis, D., Mitchell, A. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Rapid+hypothesis+testing+with+Candida+albicans+through+gene+disruption+with+short+homology+regions.">Rapid hypothesis testing with Candida albicans through gene disruption with short homology regions.</a> J. Bacteriol. 181 (6), 1868-1874 (1999).</li><li>Pulver, 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=Rsr1+focuses+Cdc42+activity+at+hyphal+tips+and+promotes+maintenance+of+hyphal+development+in+Candida+albicans.">Rsr1 focuses Cdc42 activity at hyphal tips and promotes maintenance of hyphal development in Candida albicans.</a> Eukaryotic Cell. 12 (4), 482-495 (2013).</li><li>Falgier, C., 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=Candida+species+differ+in+their+interactions+with+immature+human+gastrointestinal+epithelial+cells.">Candida species differ in their interactions with immature human gastrointestinal epithelial cells.</a> Pediatr. Res. 69 (5), 384-389 (2011).</li><li>Nosek, J., 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=Genetic+manipulation+of+the+pathogenic+yeast+Candida+parapsilosis.">Genetic manipulation of the pathogenic yeast Candida parapsilosis.</a> Curr. Genet. 42 (1), 27-35 (2002).</li><li>Zemanova, J., Nosek, J., Tomaska, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=High-efficiency+transformation+of+the+pathogenic+yeast+Candida+parapsilosis.">High-efficiency transformation of the pathogenic yeast Candida parapsilosis.</a> Curr. Genet. 45 (3), 183-186 (2004).</li><li>Benjamin, D. 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=Neonatal+candidiasis+among+extremely+low+birth+weight+infants:+risk+factors,+mortality+rates,+and+neurodevelopmental+outcomes+at+18+to+22+months.">Neonatal candidiasis among extremely low birth weight infants: risk factors, mortality rates, and neurodevelopmental outcomes at 18 to 22 months.</a> Pediatrics. 117 (1), 84-92 (2006).</li><li>Kullberg, B. J., Arendrup, M. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Invasive+Candidiasis.">Invasive Candidiasis.</a> N Engl J Med. 373 (15), 1445-1456 (2015).</li></ol>

表位的结构标记在使用上述的PCR介导的基因修饰策略念珠菌物种的序列可以归纳为一个三步过程。首先，盒通过PCR编码既期望集成和区域同源的插入轨迹入酵母基因组中的序列进行。第二，要转化的酵母细胞制成用乙酸锂和共同培养与盒化学感受。第三，将细胞铺于选择性培养基以回收转化体和所得菌落为正确整合盒成所需基因组基因座的测试。 有此协议内的几个关?...

念珠菌是殖民所有人类的肠道和泌尿生殖道共生真菌。下免疫缺陷的情况下，如发生与早产或从癌症治疗的免疫抑制作用， 念珠菌物种可以成为机会致病菌。 念珠菌物种的， 白色念珠菌是最普遍的真菌殖民者并且使得大多数侵袭性真菌感染。其它念珠菌属种类，如光滑念珠菌 ， 近平滑念珠菌 ， 热带念珠菌和C. kruseii也造成免疫功能低下患者的严重感染，有些显示出固有电阻对常用抗真菌抗生素如氟康唑和两性霉素B因此，与这些物种的感染正在观察更频繁，特别是在被用抗真菌剂预防性治疗的患者。即使有适当和及时的一NTI真菌治疗，侵袭性念珠菌感染继续与显著的发病率和死亡率的1个相关。因为在人体健康念珠菌属种类的意义，有必要为现成的分子工具，允许它们的发病机理的研究和澄清。 其中一个重要的工具，它使研究人员能够可视化和量化微生物细胞，他们表达的蛋白质是FP融合技术。聚合酶链反应（PCR）介导的基因修饰，如在本文中所描述，使融合的构建，FP序列和念珠菌蛋白在其基因组基因座编码的感兴趣序列之间。构建体的稳定整合促进蛋白表达的分析以及蛋白质定位动力学。含有FP序列的质粒，在白色念珠菌中表达优化的，所用的PCR介导的用于g烯改性的策略，已经预先构造2，3，4，5。质粒含有FP转换“盒”：链接到有利于白色念珠菌和近平滑念珠菌 2，3，4，5，6，7的转化的营养标记基因的FP序列。目前可获得的质粒含有多种对营养缺陷型菌株的转化选择性营养标记基因（URA3，HIS1，ARG4）以及显性药物抗性标记（NAT1），这有利于缺乏营养缺陷型临床株的转化。此外，质粒含有多达四个不同的FP序列（绿色[GFP]选项黄釉W [YFP]，青色[CFP]和樱桃[mCherry]）和任一用于施工羧基末端蛋白融合物，或用于建筑氨基末端蛋白质融合的启动子序列的ADH1终止序列。引物设计有同源性的周围在FP带盒的质粒DNA。此外，引物还包含轴承的同源性的感兴趣的酵母基因进行标记，这有利于带盒的整合到通过同源重组（ 图1）的基因组基因座5&#39;延伸序列。基因特异性FP盒通过PCR生成，然后转化到通过用乙酸锂处理而处于感受态用于DNA的摄取念珠菌细胞。 <img alt="图1" src="/files/ftp_upload/55333/55333fig1.jpg" /> 图1：FP序列融合如何在念珠菌属种类产生的图。 （A）质粒DNA大型的源码ES一个FP序列和编码序列诺尔丝菌素抗性（NAT1）。正向（FWD）和反向（REV）的引物的相对位置示出，以指示同源的区域与质粒序列的引物和表示特定基因同源区或引物延伸的紫色部分的黑色部分。 （B）中的FP盒被变换成念珠菌和通过同源重组（虚线）的ENO1基因座内集成。 （C）在ENO1的3&#39;末端所得的FP融合序列。 <a href="http://ecsource.jove.com/files/ftp_upload/55333/55333fig1large.jpg" target="_blank">请点击此处查看该图的放大版本。</a> 在此，我们提出在念珠菌蛋白融合物（左心室Eno1-FP）的结构的一个例子。我们使用含标记与编码GFP，YFP序列沿着NAT1转化标记基因的质粒，或mCherry（ 图2）。将这些质粒与在PCR引物一起使用，以产生特定基因盒便于FP的融合ENO1的3&#39;末端，从而导致在其羧基末端融合到的FP左心室Eno1的表达。 <img alt="图2" src="/files/ftp_upload/55333/55333fig2.jpg" /> 图2：FP含盒式质粒的图谱。向前（F）和用于产生从所述质粒卡匣倒车档（R）的引物与它们的同源性的质粒的相对位置沿指示。引物序列如表1所列。 F1和R1也被用来生成所述pYFP- NAT1盒。含有YFP- NAT1盒（pMG2263）质粒是与YFP的代替GFP序列的异常相同pMG2120。盒尺寸：GFP-NAT1，3.7 KBP; mCherry- NAT1，3.2 KBP; YFP- NAT1，3。7 KBP。这个数字已经从Gerami -内贾德， 等修改。 4 <a href="http://ecsource.jove.com/files/ftp_upload/55333/55333fig2large.jpg" target="_blank">请点击此处查看该图的放大版本。</a>

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>100W mercury lamp</td>
			<td>CHIU Technical Corporation</td>
			<td>M-100T</td>
			<td></td>
		</tr>
		<tr>
			<td>95% Ethanol</td>
			<td>Any</td>
			<td>NA</td>
			<td></td>
		</tr>
		<tr>
			<td>Adenine</td>
			<td>Any</td>
			<td>NA</td>
			<td></td>
		</tr>
		<tr>
			<td>Ampicillin</td>
			<td>Any</td>
			<td>NA</td>
			<td></td>
		</tr>
		<tr>
			<td>Carrier DNA</td>
			<td>Ambion</td>
			<td>AM9680</td>
			<td>Sheared Salmon Sperm DNA 10 mg/ml</td>
		</tr>
		<tr>
			<td>CCD Camera</td>
			<td>Photometrics</td>
			<td>CoolSNAP HQ</td>
			<td></td>
		</tr>
		<tr>
			<td>Conical Tube</td>
			<td>Corning</td>
			<td>430828</td>
			<td>50ml</td>
		</tr>
		<tr>
			<td>Culture Tube Rotator</td>
			<td>New Brunswick</td>
			<td>2013923</td>
			<td>TC-8, or Any Culture Tube Rotator</td>
		</tr>
		<tr>
			<td>Deoxynucleotides (dNTP) PCR Grade</td>
			<td>Any</td>
			<td>NA</td>
			<td></td>
		</tr>
		<tr>
			<td>Eppendorf Tubes</td>
			<td>Eppendorf</td>
			<td>022363719, 022363212</td>
			<td>0.5ml, 1.5ml</td>
		</tr>
		<tr>
			<td>Erlenmeyer Flask</td>
			<td>Fisher Scientific</td>
			<td>7250089</td>
			<td>125ml</td>
		</tr>
		<tr>
			<td>Ethylenediaminetetraacetic Acid (EDTA)</td>
			<td>Any</td>
			<td>NA</td>
			<td></td>
		</tr>
		<tr>
			<td>Freezer (-80 &#176;C )</td>
			<td>Thermo Electron Corporation</td>
			<td>ULT-1386-9-V</td>
			<td>Revco Ultima II</td>
		</tr>
		<tr>
			<td>GFP, YFP and Texas Red Filter Sets</td>
			<td>Chroma Technology Corporation</td>
			<td>49002, 86004v2, 49008</td>
			<td></td>
		</tr>
		<tr>
			<td>Glass culture tubes</td>
			<td>Fisher Scientific</td>
			<td>1496126</td>
			<td>75mm</td>
		</tr>
		<tr>
			<td>HRP goat anti-mouse antibody</td>
			<td>Santa Cruz Biotechnology</td>
			<td>SC-2005</td>
			<td></td>
		</tr>
		<tr>
			<td>HRP goat anti-rabbit antibody</td>
			<td>Santa Cruz Biotechnology</td>
			<td>SC-2301</td>
			<td></td>
		</tr>
		<tr>
			<td>Incubator (30 &#176;C )</td>
			<td>Any</td>
			<td>NA</td>
			<td></td>
		</tr>
		<tr>
			<td>Lithium Acetate</td>
			<td>Any</td>
			<td>NA</td>
			<td></td>
		</tr>
		<tr>
			<td>Lysogeny Broth (LB) Media</td>
			<td>Any</td>
			<td>NA</td>
			<td></td>
		</tr>
		<tr>
			<td>Magnesium Chloride</td>
			<td>Any</td>
			<td>NA</td>
			<td></td>
		</tr>
		<tr>
			<td>Microcentrifuge</td>
			<td>Eppendorf</td>
			<td>5415 D</td>
			<td></td>
		</tr>
		<tr>
			<td>Microscope</td>
			<td>Nikon</td>
			<td>E600</td>
			<td>Nikon Eclipse E600</td>
		</tr>
		<tr>
			<td>Microscope Image Analysis Software</td>
			<td>Universal Imaging Corporation</td>
			<td>6.3r7</td>
			<td>MetaMorph Software Series 6.3r7</td>
		</tr>
		<tr>
			<td>Mouse anti-GFP antibody</td>
			<td>Roche</td>
			<td>11814460001</td>
			<td></td>
		</tr>
		<tr>
			<td>Nourseothricin</td>
			<td>Fisher Scientific</td>
			<td>50997939</td>
			<td></td>
		</tr>
		<tr>
			<td>PCR Thermocycler</td>
			<td>Applied Biosystems</td>
			<td>9700</td>
			<td>GeneAmp PCR System</td>
		</tr>
		<tr>
			<td>PCR tubes</td>
			<td>BioExpress, GeneMate</td>
			<td>T-3035-1</td>
			<td>0.2ml</td>
		</tr>
		<tr>
			<td>Polyethylene Glycol 3350</td>
			<td>Any</td>
			<td>NA</td>
			<td></td>
		</tr>
		<tr>
			<td>Potassium Chloride</td>
			<td>Any</td>
			<td>NA</td>
			<td></td>
		</tr>
		<tr>
			<td>Rabbit anti-mCherry antibody</td>
			<td>BioVision</td>
			<td>5993-100</td>
			<td></td>
		</tr>
		<tr>
			<td>Refrigerator (4&#176;C)</td>
			<td>Any</td>
			<td>NA</td>
			<td></td>
		</tr>
		<tr>
			<td>Sodium Acetate</td>
			<td>Any</td>
			<td>NA</td>
			<td></td>
		</tr>
		<tr>
			<td>Stereomicroscope</td>
			<td>Nikon</td>
			<td>SMZ1500</td>
			<td></td>
		</tr>
		<tr>
			<td>Table Top Centrifuge</td>
			<td>Labnet</td>
			<td>Z 400</td>
			<td>Hermle Z 400</td>
		</tr>
		<tr>
			<td>Taq DNA Polymerase</td>
			<td>Any</td>
			<td>NA</td>
			<td></td>
		</tr>
		<tr>
			<td>Tris(hydroxymethyl)aminomethane (Tris)</td>
			<td>Any</td>
			<td>NA</td>
			<td></td>
		</tr>
		<tr>
			<td>Vortex Mixer</td>
			<td>Scientific Industries</td>
			<td>SI-0236</td>
			<td>Vortex Genie 2</td>
		</tr>
		<tr>
			<td>Yeast Extract Peptone Dextrose (YPD) Media</td>
			<td>Any</td>
			<td>NA</td>
			<td></td>
		</tr>
	</tbody>
</table>

generation of fluorescent protein fusions in candida species

念珠菌属种类，肠道和泌尿生殖道的常见殖民者，是多数人类侵袭性真菌感染的原因。因此，需要分子和遗传工具，以促进它们的发病机理的研究。 PCR介导的基因修饰是产生表位标记的蛋白质以促进其检测一个简单，快捷的方式。特别是，荧光蛋白（FP）融合体是功能强大的工具，使酵母细胞和蛋白质的可视化和定量分别由荧光显微镜和免疫印迹。含FP编码序列，有利于念珠菌的改造营养标记基因的质粒一起，已经为FP建设和表达念珠菌的目的产生的。在此，我们提出了在念珠菌构建FP融合的战略。包含诺尔丝菌素resista质粒NCE变换标记基因（NAT1）与绿色，黄色，或樱桃的FP（GFP，YFP，mCherry）与引物包括在聚合酶链反应（PCR）的基因特异性序列一起使用，以产生一个FP盒序列沿。这个特定基因盒具有集成到通过同源重组的相应基因座的3&#39;端的能力。成功的框内融合将FP序列插入的感兴趣的基因位点的是基因验证，然后通过显微镜和/或免疫检测方法的融合蛋白表达的分析。此外，对于高表达的蛋白质的情况下，成功的融合可以通过荧光成像技术筛选为主。

作为一个例子，我们使用上述的协议来构建GFP和mCherry融合在一个近平滑念珠菌实验室菌株ENO1。每个推定转化最初是重新划线增长。在本实施例中，由于所得到的融合蛋白中高度表达（烯醇）和FP的明亮，我们能够进行诊断性PCR之前筛选用荧光显微镜转化体（ 图3）6。 <img alt="图3" src="/files/f...

Watch this Scientific Journal Video about Generation of Fluorescent Protein Fusions in Candida Species at JoVE.com

Generation of Fluorescent Protein Fusions in Candida Species

PCR介导的基因修饰可用于产生在念珠菌属种类，这有利于可视化和酵母细胞和蛋白质的定量荧光蛋白融合体。在此，我们提出了在近平滑念珠菌构建荧光蛋白融合（左心室Eno1-FP）的战略。

在荧光蛋白融合的一代<em&gt;念珠菌</em&gt;物种

Candida species, prevalent colonizers of the intestinal and genitourinary tracts, are the cause of the majority of invasive fungal infections in humans. ...

Research

JoVE Journal

Genetics

Generation of Fluorescent Protein Fusions in Candida Species

High Resolution Fluorescent In Situ Hybridization in Drosophila Embryos and Tissues Using Tyramide Signal Amplification

High Resolution Fluorescent In Situ Hybridization in Drosophila Embryos and Tissues Using Tyramide Signal Amplification

高分辨率荧光原位杂交在果蝇胚胎和组织中使用 Tyramide 信号放大

In our efforts to determine the patterns of expression and subcellular localization of Drosophila RNAs on a genome-wide basis, and in a variety of ...

科研

遗传学

Targeted Next-generation Sequencing and Bioinformatics Pipeline to Evaluate Genetic Determinants of Constitutional Disease

有针对性的下一代测序和生物信息学管道评价宪政疾病的遗传决定因素

Next-generation sequencing (NGS) is quickly revolutionizing how research into the genetic determinants of constitutional disease is performed. The ...

CRISPR-mediated Genome Editing of the Human Fungal Pathogen Candida albicans

CRISPR-mediated Genome Editing of the Human Fungal Pathogen Candida albicans

crispr 介导的人类真菌病原体白色念珠菌基因组编辑

This method describes the efficient CRISPR mediated genome editing of the diploid human fungal pathogen Candida albicans. CRISPR-mediated genome editing ...

Use of Single Molecule Fluorescent In Situ Hybridization (SM-FISH) to Quantify and Localize mRNAs in Murine Oocytes

Use of Single Molecule Fluorescent In Situ Hybridization SM-FISH to Quantify and Localize mRNAs in Murine Oocytes

单分子荧光原位杂交 (SM-FISH) 在小鼠卵母细胞中 Mrna 的定量和定位

Current methods routinely used to quantify mRNA in oocytes and embryos include digital reverse-transcription polymerase chain reaction (dPCR), ...

Mass Spectrometry-Based Proteomics Analyses Using the OpenProt Database to Unveil Novel Proteins Translated from Non-Canonical Open Reading Frames

基于质谱的蛋白质组学分析, 利用 OpenProt 数据库揭开从非规范开放阅读框架翻译的新蛋白质

Genome annotation is central to today's proteomic research as it draws the outlines of the proteomic landscape. Traditional models of open reading frame ...

A Bioinformatics Pipeline to Accurately and Efficiently Analyze the MicroRNA Transcriptomes in Plants

生物信息学管道，用于准确、高效地分析植物中的微RNA转录

MicroRNAs (miRNAs) are 20- to 24-nucleotide (nt) endogenous small RNAs (sRNAs) extensively existing in plants and animals that play potent roles in ...

Genetic Mapping of Thermotolerance Differences Between Species of Saccharomyces Yeast via Genome-Wide Reciprocal Hemizygosity Analysis

Genetic Mapping of Thermotolerance Differences Between Species of Saccharomyces Yeast via Genome-Wide Reciprocal Hemizygosity Analysis

通过全基因组互惠性分析,对糖母菌菌种间热耐性差异的遗传图谱

A central goal of modern genetics is to understand how and why organisms in the wild differ in phenotype. To date, the field has advanced largely on the ...

Generation of Defined Genomic Modifications Using CRISPR-CAS9 in Human Pluripotent Stem Cells

在人类多能干细胞中使用CRISPR-CAS9生成定义的基因组修饰

Human pluripotent stem cells offer a powerful system to study gene function and model specific mutations relevant to disease. The generation of precise ...

Dissection of Drosophila melanogaster Flight Muscles for Omics Approaches

Dissection of Drosophila melanogaster Flight Muscles for Omics Approaches

解剖果蝇黑色素组飞行肌肉为奥米克斯方法

Drosophila flight muscle is a powerful model to study diverse processes such as transcriptional regulation, alternative splicing, metabolism, and ...

Application of CRISPR Interference (CRISPRi) for Gene Silencing in Pathogenic Species of Leptospira

Application of CRISPR Interference CRISPRi for Gene Silencing in Pathogenic Species of Leptospira

CRISPR干扰（CRISPRI）在瘦素病原物种中基因沉默的应用

Leptospirosis is a global neglected zoonosis, responsible for at least 1 million cases per year and almost 60 thousand deaths. The disease is caused by ...