{{'Search' | translate}}
 

QIAGEN Proteinase K

蛋白酶K.

Company: QIAGEN
Catalog#: 19131
Bio-protocol()
Company-protocol()
Other protocol()

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 ...

Telomere Restriction Fragment (TRF) Analysis
Author:
Date:
2015-11-20
[Abstract]  While telomerase is expressed in ~90% of primary human tumors, most somatic tissue cells except transiently proliferating stem-like cells do not have detectable telomerase activity (Shay and Wright, 1996; Shay and Wright, 2001). Telomeres progressively shorten with each cell division in normal cells, including proliferating stem-like cells, due to the end replication (lagging strand synthesis) problem and other causes such as oxidative damage, therefore all somatic cells have limited cell proliferation capacity (Hayflick limit) (Hayflick and Moorhead, 1961; Olovnikov, 1973). The progressive telomere shortening eventually leads to growth arrest in normal cells, which is known as replicative senescence (Shay et al., 1991). Once telomerase is activated in cancer cells, telomere ... [摘要]  虽然端粒酶在约90%的原发性人类肿瘤中表达,但除了瞬时增殖的干细胞样细胞之外,大多数体细胞组织细胞不具有可检测的端粒酶活性(Shay和Wright,1996; Shay和Wright,2001)。由于末端复制(滞后链合成)问题和其它原因例如氧化损伤,端粒在正常细胞中的每个细胞分裂(包括增殖的干细胞样细胞)逐渐缩短,因此所有体细胞具有有限的细胞增殖能力(Hayflick极限) (Hayflick和Moorhead,1961; Olovnikov,1973)。渐进性端粒缩短最终导致正常细胞中的生长停滞,其被称为复制衰老(Shay等人,1991)。一旦端粒酶在癌细胞中被激活,通过在染色体末端添加TTAGGG重复来稳定端粒长度,从而使细胞分裂无限延续(Shay和Wright,1996; Shay和Wright,2001)。因此,衰老和癌症之间的联系可以部分地解释端粒生物学。有许多快速和方便的方法来研究端粒生物学,例如端粒限制性片段(TRF),端粒重复扩增方案(Telomere Repeat Amplification Protocol, TRAP)(Mender and Shay,2015b)和端粒功能障碍诱导Foci(TIF)分析 ...

Yeast DNA Replication 2D Gel Protocol
Author:
Date:
2012-07-05
[Abstract]  Two-dimensional agarose gel electrophoresis (2D gel) analysis is used extensively as a method to detect origins of replication. Here, I present a simplified method for the isolation of yeast genomic DNA for 2D gel analysis from a small number of yeast cells. This DNA isolation method is simpler and less time consuming than the traditional method that involves CsCl density gradient centrifugation. This method could be modified for 2D gel analysis in other organisms as well. [摘要]  二维琼脂糖凝胶电泳(2D凝胶)分析广泛用作检测复制起点的方法。 在这里,我提出了一个简单的方法分离酵母基因组DNA的2D凝胶分析从少量的酵母细胞。 这种DNA分离方法比涉及CsCl密度梯度离心的传统方法更简单和更省时。 这种方法可以修改为其他生物体的2D凝胶分析。

Comments