Name: a cell free assay to study chromatin decondensation at the end of mitosis
Uploaded: 2015-12-19T13:00:00
Description: Watch this Scientific Journal Video about A Cell Free Assay to Study Chromatin Decondensation at the End of Mitosis at JoVE.com

这项工作得到了德国研究基金会和ERC（AN377 / 3-2和309528 CHROMDECON到华盛顿州）和勃林格殷格翰全宗的博士研究生奖学金，以AKS图1和2是从的Developmental Cell 31（3），Magalska转载等人 ，RuvB样ATP酶在有丝分裂，305-318，2014年年底的功能染色质解聚，与爱思唯尔许可。

从HeLa细胞有丝分裂染色质集群隔离 1.准备<ol><li> 细胞培养液 <ol><li>通过加入10％胎牛血清，100单位/毫升青霉素，100微克/毫升链霉素和2mM谷氨酰胺的DMEM中制备完整的Dulbecco改进的Eagle培养基（DMEM）中。制备含有2.7毫米氯化钾，137mM的氯化钠，10mM的磷酸氢二钠2H 2 O和2mM KH 2 PO 4在去离子水中，并调节pH至7.4用10N NaOH磷...

<ol>
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J., Heald, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Methods+for+studying+spindle+assembly+and+chromosome+condensation+in+Xenopus+egg+extracts.">Methods for studying spindle assembly and chromosome condensation in Xenopus egg extracts.</a> Methods Mol Biol. 322, 459-474 (2006).</li><li>Galy, V., 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+role+for+gp210+in+mitotic+nuclear-envelope+breakdown.">A role for gp210 in mitotic nuclear-envelope breakdown.</a> J Cell Sci. 121, 317-328 (2008).</li><li>Chan, R. C., Forbes, D. I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=In+vitro+study+of+nuclear+assembly+and+nuclear+import+using+Xenopus+egg+extracts.">In vitro study of nuclear assembly and nuclear import using Xenopus egg extracts.</a> Methods Mol Biol. 322, 289-300 (2006).</li><li>Gillespie, P. J., Gambus, A., Blow, J. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Preparation+and+use+of+Xenopus+egg+extracts+to+study+DNA+replication+and+chromatin+associated+proteins.">Preparation and use of Xenopus egg extracts to study DNA replication and chromatin associated proteins.</a> Methods. 57, 203-213 (2012).</li><li>Gant, T. M., Wilson, K. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Nuclear+assembly.">Nuclear assembly.</a> Annu Rev Cell Dev Biol. 13, 669-695 (1997).</li><li>Magalska, 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=RuvB-like+ATPases+function+in+chromatin+decondensation+at+the+end+of+mitosis.">RuvB-like ATPases function in chromatin decondensation at the end of mitosis.</a> Developmental Cell. 31, 305-318 (2014).</li><li>Belmont, A. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mitotic+chromosome+structure+and+condensation.">Mitotic chromosome structure and condensation.</a> Curr Opin Cell Biol. 18, 632-638 (2006).</li><li>Lavoie, B. D., Tuffo, K. M., Oh, S., Koshland, D., Holm, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mitotic+chromosome+condensation+requires+Brn1p,+the+yeast+homologue+of+Barren.">Mitotic chromosome condensation requires Brn1p, the yeast homologue of Barren.</a> Mol Biol Cell. 11, 1293-1304 (2000).</li><li>Philpott, A., Leno, G. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Nucleoplasmin+remodels+sperm+chromatin+in+Xenopus+egg+extracts.">Nucleoplasmin remodels sperm chromatin in Xenopus egg extracts.</a> Cell. 69, 759-767 (1992).</li><li>Philpott, A., Leno, G. H., Laskey, R. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Sperm+decondensation+in+Xenopus+egg+cytoplasm+is+mediated+by+nucleoplasmin.">Sperm decondensation in Xenopus egg cytoplasm is mediated by nucleoplasmin.</a> Cell. 65, 569-578 (1991).</li><li>Burglin, T. R., Mattaj, I. W., Newmeyer, D. D., Zeller, R., De Robertis, E. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cloning+of+nucleoplasmin+from+Xenopus+laevis+oocytes+and+analysis+of+its+developmental+expression.">Cloning of nucleoplasmin from Xenopus laevis oocytes and analysis of its developmental expression.</a> Genes Dev. 1, 97-107 (1987).</li><li>Whitaker, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Calcium+at+fertilization+and+in+early+development.">Calcium at fertilization and in early development.</a> Physiological reviews. 86, 25-88 (2006).</li><li>Gasser, S. M., Laemmli, U. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Improved+methods+for+the+isolation+of+individual+and+clustered+mitotic+chromosomes.">Improved methods for the isolation of individual and clustered mitotic chromosomes.</a> Exp Cell Res. 173, 85-98 (1987).</li><li>Eisenhardt, N., Schooley, A., Antonin, W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Xenopus+in+vitro+assays+to+analyze+the+function+of+transmembrane+nucleoporins+and+targeting+of+inner+nuclear+membrane+proteins.">Xenopus in vitro assays to analyze the function of transmembrane nucleoporins and targeting of inner nuclear membrane proteins.</a> Methods Cell Biol. 122, 193-218 (2014).</li><li>Wignall, S. M., Deehan, R., Maresca, T. J., Heald, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+condensin+complex+is+required+for+proper+spindle+assembly+and+chromosome+segregation+in+Xenopus+egg+extracts.">The condensin complex is required for proper spindle assembly and chromosome segregation in Xenopus egg extracts.</a> J Cell Biol. 161, 1041-1051 (2003).</li><li>Forbes, D. J., Kirschner, M. W., Newport, J. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Spontaneous+formation+of+nucleus-like+structures+around+bacteriophage+DNA+microinjected+into+Xenopus+eggs.">Spontaneous formation of nucleus-like structures around bacteriophage DNA microinjected into Xenopus eggs.</a> Cell. 34, 13-23 (1983).</li><li>Hartl, P., Olson, E., Dang, T., Forbes, D. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Nuclear+assembly+with+lambda+DNA+in+fractionated+Xenopus+egg+extracts:+an+unexpected+role+for+glycogen+in+formation+of+a+higher+order+chromatin+intermediate.">Nuclear assembly with lambda DNA in fractionated Xenopus egg extracts: an unexpected role for glycogen in formation of a higher order chromatin intermediate.</a> J Cell Biol. 124, 235-248 (1994).</li><li>Sandaltzopoulos, R., Blank, T., Becker, P. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Transcriptional+repression+by+nucleosomes+but+not+H1+in+reconstituted+preblastoderm+Drosophila+chromatin.">Transcriptional repression by nucleosomes but not H1 in reconstituted preblastoderm Drosophila chromatin.</a> EMBO J. 13, 373-379 (1994).</li><li>Ulbert, S., Platani, M., Boue, S., Mattaj, I. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Direct+membrane+protein-DNA+interactions+required+early+in+nuclear+envelope+assembly.">Direct membrane protein-DNA interactions required early in nuclear envelope assembly.</a> J Cell Biol. 173, 469-476 (2006).</li><li>Zhang, C., Clarke, P. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Chromatin-independent+nuclear+envelope+assembly+induced+by+Ran+GTPase+in+Xenopus+egg+extracts.">Chromatin-independent nuclear envelope assembly induced by Ran GTPase in Xenopus egg extracts.</a> Science. 288, 1429-1432 (2000).</li><li>Paulson, J. 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非洲爪蟾卵提取物是忠实地再现体外细胞过程一个非常有用的工具，并且该系 ​​统被成功地在细胞周期和细胞分裂事件2，3,5,6,17表征使用。由于隔离在卵子发生在卵核部件的大型商店，卵提取物的细胞成分的极好来源。相对于其他方法，如RNA干扰对哺乳动物组织的细胞系或遗传操作，它提供了几个优点：无细胞系统允许研究细胞过程中细胞生存力将是否则限制。?...

非洲爪蟾卵提取物是一个强大的和广泛应用的工具，研究在无细胞的测定法的简单复杂细胞事件。由于通过Lohka＆升井1首次描述它们已经被广泛地用于研究的有丝分裂过程，如染色质缩合2，主轴组件3，核膜破裂4，而且核质运输5或DNA复制6。发生在有丝分裂的末端的事件，所必需的interphasic核的改造如核膜改革与核孔复合重组小得多理解相比早期有丝分裂事件，但可使用爪蟾卵提取物7被类似研究。最近，我们建立了基于爪蟾卵提取物研究染色质解聚在有丝分裂8月底的检测，欠调查过程，等待着我TS细致的刻画。 在后生动物，染色质是高度会聚在进入有丝分裂，以便执行的遗传物质的忠实偏析。以确保该染色质是用于基因表达和DNA复制访问间期，它需要被解压缩在有丝分裂的末端。在脊椎动物中，染色质是高达50倍相比相间9有丝分裂期间更紧密，而相比之下，酵母，其中有丝分裂压实通常要低得多， 例如，只有两个倍S.酵母 10。脊椎动物的染色质解聚已大部分研究精子DNA重组的卵子受精后的环境。的分子机制，其中nucleoplasmin，一种丰富的卵母细胞的蛋白质，交换精子特异性精蛋白组蛋白H2A和H2B存储在蛋。这个过程也阐明用爪蟾卵提取11，12。然而，表达nucleoplasmin的仅限于卵母细胞13和有丝分裂的染色体不包含这些精子特异性鱼精蛋白。因此，在有丝分裂的结束染色质解聚为nucleoplasmin 独立的8。 对于体外解聚反应，我们采用的激活爪蟾卵和染色质集群从同步的HeLa细胞中分离产生的提取物。鸡蛋与钙离子载体治疗模仿的钙释放到受精过程中精子的进入所产生的卵母细胞。钙波触发细胞周期的恢复和卵子在减数分裂的第二个中期逮捕，发展到第一相间14。因此，准备形式激活卵卵提取物所代表的有丝分裂退出/间期状态，并有能力来诱导特异性的有丝分裂的退出状染色质解聚，核膜事件和孔复合体改造。对于有丝分裂通道的隔离romatin簇我们使用加塞＆的Laemmli 15，其中染色体簇从HeLa细胞中有丝分裂的同步和通过梯度离心分离在含有多胺缓冲器释放裂解发布的协议的一个略加修改的版本。

<table>
	<tbody>
		<tr>
			<td>spermine tetrahydrochloride</td>
			<td>Fluka analytical</td>
			<td>85610-25G</td>
			<td></td>
		</tr>
		<tr>
			<td>spermidine trihydrochloride</td>
			<td>Sigma&#160;</td>
			<td>S2501-5G</td>
			<td></td>
		</tr>
		<tr>
			<td>high-purity digitonin</td>
			<td>Millipore</td>
			<td>300410-1GM</td>
			<td>toxic</td>
		</tr>
		<tr>
			<td>PMSF</td>
			<td>Applichem</td>
			<td>A0999,0100</td>
			<td>toxic</td>
		</tr>
		<tr>
			<td>thymidine</td>
			<td>Calbiochem</td>
			<td>6060</td>
			<td></td>
		</tr>
		<tr>
			<td>nocodazole</td>
			<td>Calbiochem</td>
			<td>487928</td>
			<td>toxic</td>
		</tr>
		<tr>
			<td>37 % formaldehyde solution</td>
			<td>Roth</td>
			<td>7398-1</td>
			<td>toxic</td>
		</tr>
		<tr>
			<td>trypan blue solution (0.4%)</td>
			<td>Sigma</td>
			<td>T8154</td>
			<td>toxic</td>
		</tr>
		<tr>
			<td>1,4-dithiothreitol (DTT)</td>
			<td>Roth</td>
			<td>6908.2</td>
			<td></td>
		</tr>
		<tr>
			<td>AEBSF hydrochloride</td>
			<td>Applichem</td>
			<td>A1421,0001</td>
			<td></td>
		</tr>
		<tr>
			<td>pepstatin</td>
			<td>Roth</td>
			<td>2936.1/2/3</td>
			<td></td>
		</tr>
		<tr>
			<td>leupeptin</td>
			<td>Roth</td>
			<td>CN334</td>
			<td></td>
		</tr>
		<tr>
			<td>aprotinin</td>
			<td>Roth</td>
			<td>A162.3</td>
			<td></td>
		</tr>
		<tr>
			<td>Percoll (colloidal silica particles solution)</td>
			<td>GE Healthcare</td>
			<td>17-0891-01</td>
			<td></td>
		</tr>
		<tr>
			<td>glutamine</td>
			<td>Gibco</td>
			<td>25030-024</td>
			<td></td>
		</tr>
		<tr>
			<td>Penicillin-Streptomycin</td>
			<td>Gibco</td>
			<td>15140-122</td>
			<td></td>
		</tr>
		<tr>
			<td>75 cm&#178; tissue culture flasks</td>
			<td>Greiner Bio-one</td>
			<td>658175</td>
			<td></td>
		</tr>
		<tr>
			<td>heat-inactivated fetal bovine serum (FBS)</td>
			<td>Gibco</td>
			<td>10500-064</td>
			<td></td>
		</tr>
		<tr>
			<td>Homogenizer (40 mL tissue grinder)</td>
			<td>Wheaton</td>
			<td>357546</td>
			<td></td>
		</tr>
		<tr>
			<td>Neubauer chamber</td>
			<td>Assistent</td>
			<td>441/1</td>
			<td></td>
		</tr>
		<tr>
			<td>&#160;Oak Ridge Centrifuge Tubes, polycarbonate (50 ml)</td>
			<td>Nalgene</td>
			<td>3118-0050</td>
			<td></td>
		</tr>
		<tr>
			<td>100 &#181;m cell strainer, nylon</td>
			<td>BD Falcon</td>
			<td>352360</td>
			<td></td>
		</tr>
		<tr>
			<td>cytochalasin B</td>
			<td>Applichem</td>
			<td>A7657,0010</td>
			<td>toxic</td>
		</tr>
		<tr>
			<td>cycloheximide</td>
			<td>Roth</td>
			<td>8682.3</td>
			<td>toxic</td>
		</tr>
		<tr>
			<td>L-cystein</td>
			<td>Merck</td>
			<td>1,028,381,000</td>
			<td></td>
		</tr>
		<tr>
			<td>hCG available as Ovogest</td>
			<td>MSD</td>
			<td>1431593</td>
			<td></td>
		</tr>
		<tr>
			<td>PMSG available as Intergonan</td>
			<td>MSD</td>
			<td>1431015</td>
			<td></td>
		</tr>
		<tr>
			<td>A23187 (mixed calcium-magnesium-salt)</td>
			<td>Enzo</td>
			<td>ALX-450-002-M010</td>
			<td>toxic</td>
		</tr>
		<tr>
			<td>syringe needles (1.20 x 40 mm, 18 G x 1 1/2&#34;)</td>
			<td>Braun</td>
			<td>4665120</td>
			<td></td>
		</tr>
		<tr>
			<td>ATP</td>
			<td>Serva</td>
			<td>10920.03</td>
			<td></td>
		</tr>
		<tr>
			<td>GTP, 2 Na x 3 H20</td>
			<td>Roth</td>
			<td>K056.1/2/3/4</td>
			<td></td>
		</tr>
		<tr>
			<td>creatine phosphat disodium salt</td>
			<td>Calbiochem</td>
			<td>2380</td>
			<td></td>
		</tr>
		<tr>
			<td>creatine phosphokinase</td>
			<td>Sigma</td>
			<td>C3755-35KU</td>
			<td></td>
		</tr>
		<tr>
			<td>DMAP</td>
			<td>Sigma</td>
			<td>D2629-1G</td>
			<td></td>
		</tr>
		<tr>
			<td>DAPI&#160;</td>
			<td>Roche</td>
			<td>10236276001</td>
			<td></td>
		</tr>
		<tr>
			<td>PFA</td>
			<td>Sigma</td>
			<td>P-6148</td>
			<td>toxic</td>
		</tr>
		<tr>
			<td>centrifugation tubes for TLA 100 (7 x 10 mm, 5/16 x 13/16 in.)</td>
			<td>Beckman Coulter</td>
			<td>343775</td>
			<td></td>
		</tr>
		<tr>
			<td>&#34;Cell-Saver&#34; (tips with wide opening, 1000 &#181;L)</td>
			<td>Biozym</td>
			<td>729065</td>
			<td></td>
		</tr>
		<tr>
			<td>50 % glutaraldehyde solution, grade I</td>
			<td>Sigma alderich</td>
			<td>G7651-10 mL</td>
			<td>toxic</td>
		</tr>
		<tr>
			<td>0.1 % (w/v) poly-L-lysine solution</td>
			<td>Sigma</td>
			<td>P8920-100 mL</td>
			<td></td>
		</tr>
		<tr>
			<td>flat-bottom tubes (6 mL, 16.0/55 mm)</td>
			<td>Greiner Bio-one</td>
			<td>145211</td>
			<td></td>
		</tr>
		<tr>
			<td>Vectashield mounting medium</td>
			<td>Vector laboratories</td>
			<td>H1000</td>
			<td></td>
		</tr>
		<tr>
			<td>tubes for TLA120 (11 x 34 mm, 7/16 x 1 3/8 in.)</td>
			<td>Beckman Coulter</td>
			<td>343778</td>
			<td></td>
		</tr>
		<tr>
			<td>&#34;Cell-Saver&#34; (tips with wide opening, 200 &#181;L)</td>
			<td>Biozym</td>
			<td>729055</td>
			<td></td>
		</tr>
		<tr>
			<td>12 mm coverslips</td>
			<td>Thermo Scientific</td>
			<td>0784 #1</td>
			<td></td>
		</tr>
	</tbody>
</table>

a cell free assay to study chromatin decondensation at the end of mitosis

During the vertebrate cell cycle chromatin undergoes extensive structural and functional changes. Upon mitotic entry, it massively condenses into rod shaped chromosomes which are moved individually by the mitotic spindle apparatus. Mitotic chromatin condensation yields chromosomes compacted fifty-fold denser as in interphase. During exit from mitosis, chromosomes have to re-establish their functional interphase state, which is enclosed by a nuclear envelope and is competent for replication and transcription. The decondensation process is morphologically well described, but in molecular terms poorly understood: We lack knowledge about the underlying molecular events and to a large extent the factors involved as well as their regulation. We describe here a cell-free system that faithfully recapitulates chromatin decondensation in vitro, based on mitotic chromatin clusters purified from synchronized HeLa cells and X. laevis egg extract. Our cell-free system provides an important tool for further molecular characterization of chromatin decondensation and its co-ordination with processes simultaneously occurring during mitotic exit such as nuclear envelope and pore complex re-assembly.

在解聚反应的时间依赖性 图1显示了测定解聚的典型时间进程。可见的染色体在反应开始的簇decondenses并合并成一个单一的，圆和光滑的核。当卵提取物被替换蔗糖缓冲染色体簇仍稠，这表明，解聚活性存在于卵提取物。 染色质解聚是一个能源依赖的过程 在体外解聚反应可方便地操纵，例如，通过加入抑制?...

Watch this Scientific Journal Video about A Cell Free Assay to Study Chromatin Decondensation at the End of Mitosis at JoVE.com

A Cell Free Assay to Study Chromatin Decondensation at the End of Mitosis

The molecular mechanisms of the decondensation of highly compacted mitotic chromatin are ill-defined. We present a cell-free assay based on mitotic chromatin clusters isolated from HeLa cells and Xenopus laevis egg extract that faithfully reconstitutes the decondensation process in vitro.

无细胞化验，研究染色质解聚在有丝分裂的结束

During the vertebrate cell cycle chromatin undergoes extensive structural and functional changes. Upon mitotic entry, it massively condenses into rod ...

Research

JoVE Journal

Biology

Testing Visual Sensitivity to the Speed and Direction of Motion in Lizards

测试的速度和运动方向的视觉灵敏度在蜥蜴

Testing visual sensitivity in any species provides basic information regarding behaviour, evolution, and ecology. However, testing specific features of ...

科研

生物学

A Chromatin Assay for Human Brain Tissue

一个人脑组织中的染色质含量

Chronic neuropsychiatric illnesses such as schizophrenia, bipolar disease and autism are thought to result from a combination of genetic and environmental ...

Operant Learning of Drosophila at the Torque Meter

果蝇在操作性学习扭力计

For experiments at the torque meter, flies are kept on standard fly medium at 25&deg;C and 60% humidity with a 12hr light/12hr dark regime. A standardized ...

A Cell-to-cell Macromolecular Transport Assay in Planta Utilizing Biolistic Bombardment

A Cell-to-cell Macromolecular Transport Assay in Planta Utilizing Biolistic Bombardment

细胞的细胞大分子运输检测在植物利用基因枪法

Here, we present a simple and rapid protocol to detect and assess the extent of cell-to-cell macromolecular transport in planta. In this protocol, a ...

Chromatin Immunoprecipitation Assay for Tissue-specific Genes using Early-stage Mouse Embryos

使用早期的小鼠胚胎组织特异性基因的染色质免疫沉淀检测

Chromatin immunoprecipitation (ChIP) is a powerful tool to identify protein:chromatin interactions that occur in the context of living cells 1-3. This ...

Chromatin Interaction Analysis with Paired-End Tag Sequencing (ChIA-PET) for Mapping Chromatin Interactions and Understanding Transcription Regulation

Chromatin Interaction Analysis with Paired-End Tag Sequencing ChIA-PET for Mapping Chromatin Interactions and Understanding Transcription Regulation

配对末端标签测序（嘉PET）测绘染色质相互作用和了解转录调控染色质相互作用分析

Genomes are organized into three-dimensional structures, adopting higher-order conformations inside the micron-sized nuclear spaces 7, 2, 12. Such ...

Analysis of RNA Processing Reactions Using Cell Free Systems: 3' End Cleavage of Pre-mRNA Substrates in vitro

Analysis of RNA Processing Reactions Using Cell Free Systems: 3' End Cleavage of Pre-mRNA Substrates in vitro

前体mRNA底物的3&#39;末端切割：用无细胞体系中的RNA加工反应分析<em&gt;在体外</em

The 3&#8217; end of mammalian mRNAs is not formed by abrupt termination of transcription by RNA polymerase II (RNPII). Instead, RNPII synthesizes ...

In vitro Methylation Assay to Study Protein Arginine Methylation

In vitro Methylation Assay to Study Protein Arginine Methylation

在体外甲基化分析研究蛋白质精氨酸甲基化

Protein arginine methylation is one of the most abundant post-translational modifications in the nucleus. Protein arginine methylation can be identified ...

Quantitative Immunofluorescence Assay to Measure the Variation in Protein Levels at Centrosomes

定量免疫检测，以衡量在中心体的变异蛋白水平

Centrosomes are small but important organelles that serve as the poles of mitotic spindle to maintain genomic integrity or assemble primary cilia to ...

Chromatin Immunoprecipitation Assay for the Identification of Arabidopsis Protein-DNA Interactions In Vivo

Chromatin Immunoprecipitation Assay for the Identification of Arabidopsis Protein-DNA Interactions In Vivo

染色质免疫沉淀拟南芥蛋白-DNA相互作用的鉴定<em&gt;在体内</em

Intricate gene regulatory networks orchestrate biological processes and developmental transitions in plants. Selective transcriptional activation and ...