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Agilent Bioanalyzer High Sensitivity DNA chip Kit

安捷伦高灵敏度DNA试剂盒

Company: Agilent Technologies
Catalog#: 5067-4626
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Quantifying Symmetrically Methylated H4R3 on the Kaposi’s Sarcoma-associated Herpesvirus (KSHV) Genome by ChIP-Seq
Author:
Date:
2018-03-20
[Abstract]  Post-translational modifications to histone tails contribute to the three-dimensional structure of chromatin and play an important role in determining the relative expression of nearby genes. One such modification is symmetric di-methylation of arginine residues, which may exhibit different effects on gene expression including blocking the binding of transcriptional activators, or recruiting repressive effector molecules. Recent ChIP-Seq studies have demonstrated the importance of cross-talk between different histone modifications in gene regulation. Thus, to acquire a comprehensive understanding of the combined efforts of these epigenetic marks, ChIP-Seq must be utilized for identifying specific enrichment on the chromatin. Tumorigenic herpesvirus KSHV, employs epigenetic mechanisms for ... [摘要]  组蛋白尾部的翻译后修饰有助于染色质的三维结构,并在确定附近基因的相对表达中起重要作用。一种这样的修饰是精氨酸残基的对称二甲基化,其可能对基因表达展现出不同的作用,包括阻断转录激活剂的结合或招募抑制效应分子。最近的ChIP-Seq研究已经证明不同组蛋白修饰之间在基因调节中的相互作用的重要性。因此,为了全面了解这些表观遗传标记的共同努力,必须利用ChIP-Seq来鉴定染色质上的特定富集。利用表观遗传机制进行基因调控,并且通过ChIP-Seq以全面,不偏倚的方式评估多种组蛋白修饰的相对丰度,我们可以获得关于病毒复制和发病机理的复杂机制的更好的见解。

【背景】卡波西肉瘤相关疱疹病毒(KSHV)是一种致癌性人类病毒,在其生命周期中有两个不同阶段。在最初感染后,KSHV在宿主中建立持续的终生感染,这对免疫受损的个体来说特别有问题。 KSHV可引起HIV / AIDS患者中的各种肿瘤,包括卡波西肉瘤和多发性B细胞淋巴瘤(Chang等,1994; Cesarman等,1995; Russo ,1996; Soulier ,1995)。 KSHV拥有大约165,000bp的大型基因组,编码近90种不同的开放阅读框,具有逃避宿主免疫监视系统,改变宿主细胞生长途径和产生感染性后代病毒粒子的充足工具。

在潜伏阶段期间,仅有一部分病毒基因被表达,其在本质上是致癌的并且还有助于病毒附加体复制和传递到分裂的肿瘤细胞中(Uppal等人,2014; ...

Coupling Exonuclease Digestion with Selective Chemical Labeling for Base-resolution Mapping of 5-Hydroxymethylcytosine in Genomic DNA
Author:
Date:
2018-03-05
[Abstract]  This protocol is designed to obtain base-resolution information on the level of 5-hydroxymethylcytosine (5hmC) in CpGs without the need for bisulfite modification. It relies on (i) the capture of hydroxymethylated sequences by a procedure known as ‘selective chemical labeling’ (see Szulwach et al., 2012) and (ii) the digestion of the captured DNA by exonucleases. After Illumina sequencing of the digested DNA fragments, an ad hoc bioinformatic pipeline extracts the information for further downstream analysis. [摘要]  该协议旨在获得CpGs中5-羟甲基胞嘧啶(5hmC)水平的碱基分辨率信息,而无需亚硫酸氢盐修饰。 它依赖于(i)通过称为“选择性化学标记”(参见Szulwach等人,2012)的方法捕获羟甲基化序列和(ii)通过外切核酸酶消化捕获的DNA。 在消化的DNA片段的Illumina测序之后,特设的生物信息学管道提取信息用于进一步的下游分析。

【背景】基因组DNA中胞嘧啶的甲基化可以被蛋白质读取,并且主要被翻译成基因沉默。基因组中的大多数CpG二核苷酸是甲基化的,包括位于基因调控区如增强子的那些。然而,当需要时,这些CpG可以通过Ten Eleven Translocation(TET)酶将甲基氧化并且通过碱基切除修复系统用未甲基化的胞嘧啶置换来去甲基化。 5-羟甲基胞嘧啶(5hmC)是5-甲基胞嘧啶的第一个氧化衍生物,并且在基因组中绘制该修饰的碱基提供了关于正在进行活性去甲基化的区域的信息。尽管选择性化学标记(SCL)可以非常特异地检测5hmC,但该技术的分辨率受DNA片段大小的限制,特别是当捕获的DNA中存在多个CpG时。为了提高分辨率,我们引入了使用外切核酸酶的消化步骤,所述核酸外切酶将DNA分子修剪成靠近羟甲基化的胞嘧啶(Sérandour et。,2016)。然后对测序读数进行适当的生物信息学处理,然后将羟甲基化评分赋予捕获的CpG。

Adapting the Smart-seq2 Protocol for Robust Single Worm RNA-seq
Author:
Date:
2018-02-20
[Abstract]  Most nematodes are small worms that lack enough RNA for regular RNA-seq protocols without pooling hundred to thousand of individuals. We have adapted the Smart-seq2 protocol in order to sequence the transcriptome of an individual worm. While developed for individual Steinernema carpocapsae and Caenorhabditis elegans larvae as well as embryos, the protocol should be adaptable for other nematode species and small invertebrates. In addition, we describe how to analyze the RNA-seq results using the Galaxy online environment. We expect that this method will be useful for the studying gene expression variances of individual nematodes in wild type and mutant backgrounds. [摘要]  大多数线虫是小蠕虫,缺乏足够的RNA用于常规的RNA-seq协议,而没有汇集成千上万的个体。 我们已经调整了Smart-seq2协议来排序单个蠕虫的转录组。 虽然针对Steinernema carpocapsae和Caenorhabditis elegans幼虫以及胚胎开发,但该方案应该适用于其他线虫物种和小无脊椎动物。 另外,我们介绍如何使用Galaxy在线环境分析RNA-seq结果。 我们预计这种方法将有助于研究野生型和突变体背景个体线虫的基因表达差异。

【背景】低输入RNA-seq方案和扩增试剂盒,例如Smart-seq(Takara Bio,USA,Inc)和SuperAmp(Miltenyl Biotec,Inc),已经越来越多地开发和商业化,作为对低输入RNA-基于小组织,单一微生物和单细胞的seq研究。这些研究经常探索并解决特定群体(例如细胞群体,复杂组织或微生物群体)的个体中的异源基因表达。针对微生物(如线虫)的低输入RNA-seq方案的改进和适应将通过允许在单一线虫水平上分析基因表达异质性而极大地有益于线虫领域。在这里,我们已经调整了单细胞RNA-seq方案Smart-seq2(Picelli等人,2013和2014; Trombetta等人,2014),对于单线虫RNA测序。我们成功地在昆虫寄生线虫Steinernema ...

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