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胡乱走走2011银虫 (著名写手)
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[求助]
新手chip实验中的几个问题，敬请指教! 已有1人参与
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1. 2013年07月取的胚胎样品，前期处理（即甲醛固定，甘氨酸终止固定，PBS洗涤等）后，放于-80°超低温冰箱中冻存，样品还能使用吗？ 2. 将unsheared DNA 和sheared DNA进行电泳检测超声效果，没有任何条带，后来尝试将unsheared DNA 和sheared DNA解交联（加氯化钠，65°处理3h）,电泳仍没有任何条带，为什么？ 3. 有没有人用自己制作出来的多抗，或者是单抗做出Chip实验的？ 非常感谢。 |
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我们很多的客户，在使用这款产品。您可以看看是否可以解决您的问题。或可以加技术Q1951545998 MeDIP超级试剂盒 货号：A-P-1052 EpiQuik MeDIP Ultra Kit 背景资料：Core mechanisms for epigenetic alteration of genomic DNA are hypermethylation of CpG islands in specific genes and global DNA hypomethylation. Region-specific DNA methylation plays an important role in the repression of gene transcription and is mainly found in 5’-CpG-3’dinucleotides within promoters or in the first exon of genes. Global DNA hypomethylation is likely caused by methyl-deficiency due to a variety of environmental influences. It has been demonstrated that alterations in DNA methylation are associated with many diseases, especially cancer. Highly specific isolation of methylated DNA combined with next generation sequencing for genome-wide methylation analysis should provide an advantage for convenient and comprehensive identification of methylation status of normal and diseased cells, such as cancer cells [1]. Such analysis requires the isolated methylated DNA to contain minimal background in order to achieve high specificity (>98%) for reliably identifying true methylated regions. The major method for enriching methylated DNA used for genome-wide methylation profiling is methylated DNA immunoprecipitation (MeDIP) [2]. However, currently used MeDIP methods, represented by most commercially available kits, have significant weaknesses including highly non-specific enrichment (amount of enriched DNA is >75% of the amount of input DNA) [3], time consuming, labor intensive, and has low throughput. Thus, for effectively and specifically capturing methylated DNA used for next generation sequencing analysis, an ideal MeDIP method requires maximum sensitivity with minimal background levels. Epigentek’s EpiQuik™ MeDIP Ultra Kit is designed to achieve these goals by maximizing sensitivity and minimizing non-specific background signals, and is a significant improvement over previous MeDIP kits. 产品描述：This kit includes a methylated DNA control and an unmethylated DNA control, a negative control non-immune IgG, and control primers that can be used with the control DNA to demonstrate the enrichment efficacy and specificity for methylated DNA. The 5-methylcytosine monoclonal antibody provided in this kit is highly specific against methylated DNA fragments, both single and double stranded, and is not cross-reactive to hydroxymethylated and unmethylated DNA fragments. This antibody can capture >50% of DNA fragments containing as few as two 5-mCs and enriches all DNA fragments containing four or more 5-mCs. The positive control DNA containing 5-mC can be immunoprecipitated by the 5-mC antibody but not by the non-immune IgG. In this MeDIP, immunoprecipitation of 5-mC-enriched DNA fragments is processed in a microplate under optimized reaction conditions, which enables MeDIP to be completed within 3 hours with high efficiency. Immunoprecipitated methylated DNA is then cleaned, released, and eluted. Eluted DNA can be used for various downstream applications including PCR (MeDIP-PCR) and microarray (MeDIP-chip), and is especially suitable for MeDIP-seq. 产品特点： 1·Extremely fast and convenient protocol with a total procedure time (from input sample to ready-to-use methylated DNA) of less than 3 hours, which includes a minimal handling time of less than 20 minutes； 2·Optimized buffers and protocol allow minimal background by overcoming the weaknesses that cause non-specific enrichment； 3.A highly specific 5-mC monoclonal antibody included in the kit can strongly bind both single and double stranded DNA fragments containing 2 or more 5-mCs which enables highly sensitive enrichment of methylated DNA with >99% specificity. 保存建议：在接收到艾德寄出的试剂盒后请按照说明书建议，使用不同的保存条件或温度来保存试剂盒内组分。 更多产品链接：http://www.aderr.com/cn/main.php ... 07&id=30328 |
2楼2014-04-22 23:12:12