Name: an integrated platform for genome-wide mapping of chromatin states using high-throughput chip-sequencing in tumor tissues
Uploaded: 2018-04-05T13:00:00
Description: Watch this Scientific Journal Video about An Integrated Platform for Genome-wide Mapping of Chromatin States Using High-throughput ChIP-sequencing in Tumor Tissues at JoVE.com

我们感谢马库斯科伊尔, 柯蒂斯 Gumbs, SMF 核心在 MDACC 的排序支持。本文所述的工作得到了 NIH 赠款 (CA016672) 对 SMF 核心和1K99CA160578 奖 (R00CA160578) 的赠款的支持, 以 k.r。

所有临床标本都是按照机构审查委员会的指导方针获得的。
1. 缓冲准备
<ol><li>使200毫升的 TE 缓冲器 (10 毫米三盐酸, 10 毫米乙二胺乙酸 (EDTA) pH 值 8.0)。</li>
	<li>使200毫升的圣凯瑟缓冲 (10 毫米三盐酸, 10 毫米 EDTA pH 值 8.0, 140 毫米氯化钠)。</li>
	<li>使200毫升2.0 米甘氨酸 (37.52 克甘氨酸在200毫升水) 和加热它到65°c。</li>
	<li>使200毫升的5% 脱氧钠 (doc) 溶液 (10 克的 doc 在200毫升水)?...

<ol>
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本协议描述了一种完整和全面的高通量芯片序列模块, 用于在人类肿瘤组织和细胞系中染色质状态的全基因图谱。在任何芯片序列协议中, 最重要的步骤之一是抗体特异性。在这里, 此方法说明了所描述的六组蛋白修改的免疫沉淀条件, 所有这些都是芯片级的, 以前在我们和其他实验室中验证过 42, 44, 45.虽然相同的抗体浓度已成?...

大多数哺乳动物基因组 (98-99%) 由非编码序列组成, 这些 nocoding 区域包含已知参与控制基因表达和染色质组织 1, 2 的调控元素.在正常细胞中, 基因组 DNA 的特定组装进入致密的染色质结构, 对于各种 DNA 相关进程的空间组织、调控和精确定时至关重要, 这些过程3,4,5。然而, 在癌细胞中, 异常表观遗传机制的染色质修饰可能导致染色质结构的不正确组织, 包括获得调控元素、染色体循环系统和基因表达模式6, 7,8,9,10。
尽管最近取得了进展, 我们对与肿瘤进展或治疗反应相关的表观变化的理解有限。表观基因由一系列的修改组成, 包括组蛋白标记和 DNA 甲基化, 它们共同形成一个动态状态 (称为染色质状态), 它会影响基因表达网络和其他关键的过程来维持细胞身份。最近, 通过研究 H3K27Ac 配置文件11, 增强剂的改变已显示在多种恶性肿瘤中。虽然这些研究提供了对孤立的表观遗传标记的相关性的洞察力, 但100多种表观修改已被识别为12,13 , 而没有清楚地了解它们的生物学角色和相互 依赖。此外, 还有更多的可能组合模式的这些组蛋白和 DNA 的修改, 它是这些组合模式-而不是个别的修改-听写表观遗传学的状态14。因此, 在癌症进展或对治疗的反应中, 有必要确定这些染色质状态的变化.由于技术 (例如, 从少量临床材料/单细胞生成大规模数据) 和分析 (例如算法来定义, 癌症中表观基因改变的综合知识已经滞后于部分原因。组合状态) 的挑战。因此, 迫切需要强健的高通量方法来分析大量的组蛋白修饰标记, 从临床材料和易于实现的计算方法来预测组合模式, 这将有助于不同肿瘤分期和治疗耐药性相关的表观遗传状态的测定。此外, 从最近的表观基因分析研究中获得的数据15, 16, 17, 18, 19, 20, 21, 22、23正常组织和细胞系可与肿瘤染色质剖面结合, 进一步深入了解表观基因对肿瘤生物学的贡献。
染色质分析已成为识别各种染色质修改的全局绑定模式的强大工具15,24。近年来, 芯片序列已成为研究全球尺度上的 DNA-蛋白质相互作用的 "金标准"25,26,27。对于任何芯片序列实验, 有必要的关键步骤, 它的成功, 包括组织处理和解除, 确定最佳超声波条件, 确定的最佳抗体浓度的降水, 图书馆准备,后测序数据处理和下游分析。这些步骤中的每一个都包含关键的质量控制检查点, 当两者结合在一起时, 对于正确识别功能验证的潜在目标至关重要。通过这些步骤的创新, 一些先前的研究已经开发出从少量组织执行芯片或芯片序列的方法28,29,30,31,32.此外, 一些研究还提出了高通量芯片实验的协议, 其次是基于 PCR 的定量33,34。最后, 一些公开可用的芯片序列数据分析平台现在可用, 如 Easeq35和 Galaxy36。然而, 在高吞吐量的方式下, 结合计算管道来执行单标记和染色质状态分析, 一个集成的平台来执行芯片序列。
本协议描述了一个完整和全面的芯片序列工作流程, 用于全基因组在肿瘤组织和细胞系中的染色质状态图谱, 并易于遵循指导原则, 包括成功实验所需的所有步骤。通过采用以前由 Blecher Gonen et描述的高通量方法。37, 此协议可在多个样本中并行执行, 并已成功应用于癌细胞系和人类肿瘤, 如黑色素瘤、结肠、前列腺和胶质瘤多形性。我们演示了六组核心组蛋白修饰的方法, 它代表了人类黑色素瘤细胞系和肿瘤样本中表观遗传调控景观的关键组成部分。这些修改包括 H3K27ac (增强剂)、H3K4me1 (积极和有活力的增强剂)、H3K4me3 (促进者)、H3K79me2 (转录区)、H3K27me3 (polycomb 镇压) 和 H3K9me3 (异色镇压)。这些标记可以单独或组合使用, 以确定代表压制性和活动域的功能上不同的染色质状态。

<table>
	<tbody>
		<tr>
			<td>ChIP-grade H3K4me1 antibody</td>
			<td>Abcam</td>
			<td>ab8895</td>
			<td></td>
		</tr>
		<tr>
			<td>ChIP-grade H3K27ac antibody</td>
			<td>Abcam</td>
			<td>ab4729</td>
			<td></td>
		</tr>
		<tr>
			<td>ChIP-grade H3K4me3 antibody</td>
			<td>Abcam</td>
			<td>ab8580</td>
			<td></td>
		</tr>
		<tr>
			<td>ChIP-grade H3K79me2 antibody</td>
			<td>Abcam</td>
			<td>ab3594</td>
			<td></td>
		</tr>
		<tr>
			<td>ChIP-grade H3K27me3 antibody</td>
			<td>Abcam</td>
			<td>ab6002</td>
			<td></td>
		</tr>
		<tr>
			<td>ChIP-grade H3K9me3 antibody</td>
			<td>Abcam</td>
			<td>ab8898</td>
			<td></td>
		</tr>
		<tr>
			<td>1M Tris HCl, pH 8.0</td>
			<td>Teknova</td>
			<td>T1080</td>
			<td></td>
		</tr>
		<tr>
			<td>EDTA</td>
			<td>Sigma-Aldrich</td>
			<td>E9884</td>
			<td></td>
		</tr>
		<tr>
			<td>NaCl</td>
			<td>Sigma-Aldrich</td>
			<td>S7653</td>
			<td></td>
		</tr>
		<tr>
			<td>Glycine</td>
			<td>Sigma-Aldrich</td>
			<td>G8898</td>
			<td></td>
		</tr>
		<tr>
			<td>Sodium deoxycholate</td>
			<td>Sigma-Aldrich</td>
			<td>30970</td>
			<td></td>
		</tr>
		<tr>
			<td>DPBS</td>
			<td>Sigma - Life Sciences</td>
			<td>D8537-500ML</td>
			<td></td>
		</tr>
		<tr>
			<td>SDS</td>
			<td>Sigma-Aldrich</td>
			<td>74255</td>
			<td></td>
		</tr>
		<tr>
			<td>Triton-X</td>
			<td>Sigma-Aldrich</td>
			<td>X100-100ML</td>
			<td></td>
		</tr>
		<tr>
			<td>LiCl</td>
			<td>Sigma-Aldrich</td>
			<td>746460</td>
			<td></td>
		</tr>
		<tr>
			<td>NP-40</td>
			<td>Calbiochem</td>
			<td>492016-100ML</td>
			<td></td>
		</tr>
		<tr>
			<td>1% TWEEN-20</td>
			<td>Fisher Bioreagents</td>
			<td>BP337-500</td>
			<td></td>
		</tr>
		<tr>
			<td>BSA - IgG-free</td>
			<td>Sigma - Life Sciences</td>
			<td>A2058-5G</td>
			<td></td>
		</tr>
		<tr>
			<td>HBSS</td>
			<td>Gibo</td>
			<td>14025092</td>
			<td></td>
		</tr>
		<tr>
			<td>GentleMACS C tube</td>
			<td>GentleMACS</td>
			<td>120-008-466</td>
			<td>disassociation tube</td>
		</tr>
		<tr>
			<td>16% Formaldehyde</td>
			<td>Peierce</td>
			<td>28906</td>
			<td></td>
		</tr>
		<tr>
			<td>miniProtease inhibitor&#160;</td>
			<td>Roche Diagnostics</td>
			<td>11836153001</td>
			<td>protease inhibitor tablets</td>
		</tr>
		<tr>
			<td>Dynabeads Protein G&#160;</td>
			<td>Invitrogen</td>
			<td>10009D</td>
			<td></td>
		</tr>
		<tr>
			<td>Bioruptor NGS tubes 0.65 mL</td>
			<td>Diagenode</td>
			<td>C30010011</td>
			<td>sonication tubes</td>
		</tr>
		<tr>
			<td>DynaMag - 96 Side Skirted</td>
			<td>Invitrogen</td>
			<td>120.27</td>
			<td>96-well magnetic stand</td>
		</tr>
		<tr>
			<td>TE buffer</td>
			<td>Promega</td>
			<td>V6231</td>
			<td></td>
		</tr>
		<tr>
			<td>RNase A</td>
			<td>Invitrogen</td>
			<td>12091021</td>
			<td></td>
		</tr>
		<tr>
			<td>Proteinase K</td>
			<td>Invitrogen</td>
			<td>100005393</td>
			<td></td>
		</tr>
		<tr>
			<td>AMPure XP beads</td>
			<td>Beckman Coulter</td>
			<td>A63882</td>
			<td>paramagnetic beads</td>
		</tr>
		<tr>
			<td>Ethanol</td>
			<td>Sigma-Aldrich</td>
			<td>E7023</td>
			<td></td>
		</tr>
		<tr>
			<td>Qubit ds DNA High Sensitivity Assay Kit</td>
			<td>Invitrogen</td>
			<td>Q32854</td>
			<td>high sensitivity DNA reagents</td>
		</tr>
		<tr>
			<td>NEBNext Ultra II DNA Library Prep Kit</td>
			<td>New England BioLabs</td>
			<td>E7645L</td>
			<td>DNA Library Prep Kit</td>
		</tr>
		<tr>
			<td>Nuclease-free water</td>
			<td>Ambion</td>
			<td>AM9932</td>
			<td></td>
		</tr>
		<tr>
			<td>High sensitivity D1000 ScreenTape</td>
			<td>Agilent Technologies</td>
			<td>5067-5584</td>
			<td>high sensitivity DNA reagents</td>
		</tr>
		<tr>
			<td>High sensitivity D1000 reagents</td>
			<td>Agilent Technologies</td>
			<td>5067-5585</td>
			<td>high sensitivity DNA reagents</td>
		</tr>
		<tr>
			<td>Multiplex Oligos (Index primers- Set 1)</td>
			<td>New England BioLabs</td>
			<td>E7335L</td>
			<td>Multiplex Oligos&#160;</td>
		</tr>
		<tr>
			<td>Multiplex Oligos (Index primers- Set 2)</td>
			<td>New England BioLabs</td>
			<td>E7500L</td>
			<td>Multiplex Oligos&#160;</td>
		</tr>
		<tr>
			<td>TapeStation 4200</td>
			<td>Agilent Technologies</td>
			<td>G2991AA</td>
			<td>high sensitivity DNA electropherogram instrument</td>
		</tr>
		<tr>
			<td>Bioruptor Pico sonication device&#160;</td>
			<td>Diagenode</td>
			<td>B01060001</td>
			<td>water bath disruputor</td>
		</tr>
		<tr>
			<td>Mixer</td>
			<td>Nutator</td>
			<td>421105</td>
			<td></td>
		</tr>
		<tr>
			<td>Bio-Rad C1000 Touch Thermal Cycler</td>
			<td>Bio-Rad</td>
			<td>1851196</td>
			<td>PCR Thermal cycler</td>
		</tr>
		<tr>
			<td>Water Bath</td>
			<td>Fisher Scientific</td>
			<td>2322</td>
			<td></td>
		</tr>
		<tr>
			<td>Multichannel Pipet</td>
			<td>Denville</td>
			<td>1003123</td>
			<td></td>
		</tr>
		<tr>
			<td>Tube Revolver</td>
			<td>Thermo-Scientific</td>
			<td>88881001</td>
			<td></td>
		</tr>
		<tr>
			<td>96-Well Skirted Plate</td>
			<td>Eppendorf</td>
			<td>47744-110</td>
			<td></td>
		</tr>
		<tr>
			<td>Allegra X-12R Centrifuge&#160;</td>
			<td>Beckman Coulter</td>
			<td>A99464</td>
			<td>benchtop centrifuge</td>
		</tr>
		<tr>
			<td>Centrifuge 5424</td>
			<td>Eppendorf</td>
			<td>22620461</td>
			<td>tabletop centrifuge</td>
		</tr>
		<tr>
			<td>Optical tube strips (8x Strip)</td>
			<td>Agilent Technologies</td>
			<td>401428</td>
			<td></td>
		</tr>
		<tr>
			<td>Optical tube strip caps (8x strip)</td>
			<td>Agilent Technologies</td>
			<td>401425</td>
			<td></td>
		</tr>
		<tr>
			<td>Loading Tips, 10 Pk</td>
			<td>Agilent Technologies</td>
			<td>5067-5599</td>
			<td></td>
		</tr>
		<tr>
			<td>IKA MS3 vortex</td>
			<td>IKA</td>
			<td>3617000</td>
			<td>vortex</td>
		</tr>
	</tbody>
</table>

an integrated platform for genome-wide mapping of chromatin states using high-throughput chip-sequencing in tumor tissues

组蛋白修饰是表观基因的主要组成部分, 在确定相关基因座的转录状态方面起着重要的调控作用。此外, 还使用特定的修改来确定位置和身份非编码功能元素, 如增强剂。近年来, 染色质免疫沉淀其次是下一代测序 (芯片序列) 已成为一个强有力的工具, 以确定基因组范围内的个别蛋白修饰的轮廓。然而, 越来越清楚的是, 染色质修饰的组合模式, 称为染色质状态, 决定了相关基因组轨迹的特征和性质。因此, 由健壮的高通量 (HT) 方法组成的工作流, 用于分析许多组蛋白修饰标记, 以及能够处理无数芯片序列分析数据集的计算分析管线, 因此需要大量样品中 epigenomic 状态的综合测定。这里提出的 HT 芯片序列工作流包括两个模块: 1) 一个实验性的协议, 用于分析从少量肿瘤样本和细胞系的96井格式的几个组蛋白修饰;和 2) 一个计算数据分析管道, 结合现有的工具, 以计算单独的标记占用和组合染色质状态模式。这两个模块结合在一起, 便于快速、高效地处理数以百计的芯片序列样本。本文介绍的工作流用于从黑色素瘤肿瘤和细胞系中的6组蛋白标记剖面中提取染色质状态模式。总的来说, 我们提出了一个全面的芯片序列的工作流程, 可用于许多人肿瘤标本和癌细胞株, 以确定 epigenomic 畸变在各种恶性肿瘤。

该协议允许免疫沉淀从冰冻肿瘤组织和细胞系, 可以在许多样本并行使用高通量方法 ( 图1A)。染色质碎片应介于 200-1000 bps 之间, 以达到最佳免疫沉淀。我们注意到, 达到相同的剪切长度所需的时间不同的组织和细胞类型。芯片的成功, 从少量的组织依赖于保持裂解冷, 特别是在超声波期间。在进行库准备之前, 应量化纯化的芯片 DN Ashould (<s...

Watch this Scientific Journal Video about An Integrated Platform for Genome-wide Mapping of Chromatin States Using High-throughput ChIP-sequencing in Tumor Tissues at JoVE.com

An Integrated Platform for Genome-wide Mapping of Chromatin States Using High-throughput ChIP-sequencing in Tumor Tissues

肿瘤组织中高通量芯片测序的染色质状态全基因组图谱集成平台

在这里, 我们描述了一个优化的高通量芯片排序协议和计算分析管道, 以确定从冷冻肿瘤组织和细胞系的全基因组的染色质状态模式。

Histone modifications constitute a major component of the epigenome and play important regulatory roles in determining the transcriptional status of ...

The overall goal of this protocol is to unbiasedly identify combinatorial chromatin state patterns in tumor tissues and cancer cell lines. This method could help answer key questions in the cancer epigenetics field such as how a barren histone modification states could contribute to tumor genesis. The main advantage of this technique is that multiple histone modifications, as well as multiple samples could be processed both simultaneously, as well as efficiently.First, dissociate 50 milligrams of melanoma tissue manually and two milliliters of Hank's Balanced Salt Solution or HBSS, using a sterile razor blade. Mince the tissue into three or four millimeter pieces for approximately five minutes in a sterile tissue culture dish. Transfer the tissue solution to a dissociator tube and add another eighth milliliters of HBSS.Further dissociate the tissue, using a dissociator, until homogenized. Next, crosslink the tissue, adding 200 microliters of 16%formaldehyde per three milliliters of HBSS. Using a mixer, shake the mixture at 10 rpms for exactly 10 minutes at 37 degrees Celsius.After removing the sample from the incubator, add 200 microliters of tumoral glycine per three milliliters of sample and shake the mixture at 10 rpms for five minutes at 37 degrees Celsius. Spin the sample at 934 times g for five minutes at four degrees Celsius using a bench top centrifuge. When finished, remove the supernatant and add five milliliters of ice cold PBS.Then, centrifuge the sample again at 934 times g then remove the supernatant. Add 300 microliters of ChIP harvest buffer with protease inhibitors per 50 milligrams of tissue and lyse the samples for 30 minutes on ice. While the cells are lysing, turn on the water bath disruptor and associated cooling system and allow the temperature to reach four degrees Celsius.Place the sonicator tube in the water bath disruptor and sonicate the melanoma tissue for 60 cycles at 30 seconds on and 30 seconds off to obtain chromatin fragments of 200 to 600 base pairs. For eight milligrams of melanoma tissue, incubate three micrograms of each histone antibody with 20 microliters of protein g magnetic beads and 100 microliters of binding blocking buffer for two hours at four degrees Celsius with rotation, using a tube revolver. After sonication, dilute the sample five times using ChIP dilution buffer to bring the SDS concentration down to 0.1%After incubation of the antibody and protein g magnetic beads is complete, remove the ChIP dilution buffer and re suspend the beads in 240 microliters of the fresh buffer.Then, place 20 microliters aliquots of the beads into 12 separate tubes, each of which will be used for a separate tissue sample. Aliquot the sonicated material evenly to the protein g beads with specific antibody and rotate the tubes using a tube revolver overnight at four degrees Celsius. On the morning after reverse cross linking, transfer the antibody protein solution to a 96 well plate and place it on a magnetic stand.After allowing the beads to adhere for at least 30 seconds, remove the supernatant. Wash the beads five times with 150 microliters of ice cold RIPA wash buffer using a multi channel pipette. Move the magnet continuously for 30 seconds per wash, then remove the supernatant.After the washing steps, add an elution buffer master mix, containing 44 microliters of direct elution buffer, one microliter of Rnase and five microliters of protenase k per chip and input the sample for reverse cross linking. Incubate the samples using a PCR thermal cycler for four hours at 37 degrees Celsius, four hours at 50 degrees Celsius and eight to 16 hours at 65 degrees Celsius. On the next morning, place the samples back on the magnet and transfer the supernatant to a new 96 well PCR plate which contains the amino precipitated DNA.Add 2.3 x paramagnetic beads to the solution and carefully pipette up and down 25 times using a multi channel pipette. Following this, incubate at room temperature for four minutes. Then place the samples back on the magnet and incubate at room temperature for another four minutes.After discarding the supernatant, add 150 microliters of 70%ethanol to the samples and incubate at room temperature for 30 seconds without disturbing the beads. After allowing the beads to dry, remove the samples from the magnet and add 30 microliters of 10 millimolar traceal. Mix by pipetting 25 times.After incubating the samples at room temperature, place them back on the magnet and incubate at room temperature for another four minutes. Transfer 20 microliters of each sample to a new 96 well plate for library preparation. After PCR amplification, remove the samples and bring the total volume to 100 microliters using nucleus free water.Perform double sided paramagnetic size selection by first adding 55 microliters of beads to each sample and mixing 25 times with a pipette. Then incubate at room temperature for four minutes. Place the samples back on the magnet and incubate at room temperature for another four minutes to attain large fragments on the beads that are unsuitable for sequencing.After incubation, transfer the supernatant to a new 96 well PCR plate. Add another 25 microliters of paramagnetic beads and mix by pipetting 25 times and incubate at room temperature off the magnet for four minutes. After incubating at room temperature, place the samples back on the magnet and incubate at room temperature for four minutes.Then discard the supernatant. While leaving the samples on the magnet, add 150 microliters of 70%ethanol and incubate at room temperature for 30 seconds without disturbing the beads. Remove the ethanol and repeat the wash.After allowing the beads to dry, remove the samples from the magnet and add 25 microliters of 10 millimolar traceal pH 8.0. Mix by pipetting 25 times and incubate at room temperature, off the magnet for four minutes. After incubating at room temperature, place the samples back on the magnet and incubate at room temperature for four minutes.Now, quantify the libraries using a high sensitivity DNA electropherogram instrument before multiplexing to ensure the sizes are suitable for sequencing. This protocol allows immunoprecipitation of frozen tumor tissues and cell lines that can be performed on dozens of samples and parallel using a high throughput method. Chromatin fragments should range between 200 and 1000 base pairs for optimal immunoprecipitation.Purified ChIP DNA should be quantified before proceeding to library preparation. Completed libraries suitable for multiplexing and next generation sequencing should range between 200 and 600 base pairs and be devoid of any primer dimers. Upon post sequencing, there are many quality control metrics that should be met before proceeding to further steps of the pipeline, such as macs peak calling and chromHmm analysis.ChIP samples should be enriched over input DNA and each histone modification should display a distinct profile representing a specific component of the epigenetic landscape. The chrome HMM algorithm uses a Multivariate Hidden Markov Model to identify the most prominent reoccurring combinatorial and spacial chromatin patterns based on the histone modifications studied. Using these six modifications, chrome HMM identifies functionally distinct chromatin states representing both repressive and active domains, such as polycomb repression, heterochromatic repression, active transcription, and active enhancers.Once mastered, this technique can be done in three to four days, if it's performed properly. While attempting this procedure, it's important to determine optimal sonication conditions and specification for antibodies before continuing to library preparation and data analysis. For learning this procedure, other methods, such as RRS sequencing and chromosome confirmation capture can be performed to determine coalition of chromatin state parts with gene expression and higher chromatin structure.After the experiment, the technique has paved way for researchers in cancer epigenetics field to explore chromatin state patterns in patient tumor samples. After watching this video, you should have a good understanding of how to process and disassociate tumor tissues, perform chromatin immunoprecipitation, prepare libraries for nextgen sequecing, process post sequencing data and gain insights into initial steps of downstream analysis. Don't forget that when working with formaldehyde, it can be extremely hazardous and precautions, such as an appropriate fume hood, should always be taken when performing this procedure.

Research

JoVE Journal

Cancer Research

The Drosophila Imaginal Disc Tumor Model: Visualization and Quantification of Gene Expression and Tumor Invasiveness Using Genetic Mosaics

The Drosophila Imaginal Disc Tumor Model: Visualization and Quantification of Gene Expression and Tumor Invasiveness Using Genetic Mosaics

该<em&gt;果蝇</em&gt;成虫盘瘤模型：基因的表达与肿瘤侵袭性的可视化和定量遗传马赛克

Drosophila melanogaster has emerged as a powerful experimental system for functional and mechanistic studies of tumor development and progression in the ...

科研

癌症研究

The Establishment of a Lung Colonization Assay for Circulating Tumor Cell Visualization in Lung Tissues

肺定植法在肺部组织循环肿瘤细胞可视化中的建立

Metastasis is the major cause of cancer death. The role of circulating tumor cells (CTCs) in promoting cancer metastasis, in which lung colonization by ...

Genome-wide RNAi Screening to Identify Host Factors That Modulate Oncolytic Virus Therapy

全基因组的 rna 干扰筛查以确定调节溶瘤病毒治疗的宿主因素

High-throughput genome-wide RNAi (RNA interference) screening technology has been widely used for discovering host factors that impact&#160;virus ...

Native Chromatin Immunoprecipitation Using Murine Brain Tumor Neurospheres

利用小鼠脑肿瘤球的原发染色质沉淀

Epigenetic modifications may be involved in the development and progression of glioma. Changes in methylation and acetylation of promoters and regulatory ...

Generation of High-Throughput Three-Dimensional Tumor Spheroids for Drug Screening

高通量三维肿瘤球体药物筛选的生成

Cancer cells have routinely been cultured in two dimensions (2D) on a plastic surface. This technique, however, lacks the true environment a tumor mass is ...

Digital Polymerase Chain Reaction Assay for the Genetic Variation in a Sporadic Familial Adenomatous Polyposis Patient Using the Chip-in-a-tube Format

数字聚合酶链反应法测定零星家族性腺瘤性息肉病患者的基因变异

The quantitative analysis of human genetic variation is crucial for understanding the molecular characteristics of serious medical conditions, such as ...

In Vitro Assay to Study Tumor-macrophage Interaction

体外测定研究肿瘤-巨噬菌体相互作用

Tumor associated macrophages (TAMs) account for a large percentage of cells in the tumor mass for different types of cancers. Glioblastoma (GBM), a ...

Establishment and Characterization of Small Bowel Neuroendocrine Tumor Spheroids

小肠神经内分泌肿瘤类球体的建立与特征

Small bowel neuroendocrine tumors (SBNETs) are rare cancers originating from enterochromaffin cells of the gut. Research in this field has been limited ...

Genome-Wide Analysis of DNA Methylation in Gastrointestinal Cancer

胃肠癌DNA甲基化基因组全谱分析

DNA methylation is an important epigenetic change that is biologically meaningful and a frequent focus of cancer research. Genome-wide DNA methylation is ...

Patient-Derived Tumor Explants As a "Live" Preclinical Platform for Predicting Drug Resistance in Patients

患者源性肿瘤外植体作为预测患者耐药性的"实时"临床前平台

An understanding of drug resistance and the development of novel strategies to sensitize highly resistant cancers rely on the availability of suitable ...