“Non-canonical protein-DNA interactions identified by ChIP are not artifacts”: response

BMC Genomics

Correspondence

Daniel Schindler¹  Torsten Waldminghaus¹ 

[1] LOEWE-Center for Synthetic Microbiology (SYNMIKRO), Philipps-Universität Marburg, Hans-Meerwein-Str. 6, D-35043, Marburg, Germany;

关键词: ChIP-Chip;    ChIP-Seq;    False-positive;    σ;    seqA;   

DOI  :  10.1186/1471-2164-14-638

 received in 2013-06-18, accepted in 2013-09-18,  发布年份 2013

来源: Springer

【 摘 要 】

BackgroundStudies of protein association with DNA on a genome wide scale are possible through methods like ChIP-Chip or ChIP-Seq. Massive problems with false positive signals in our own experiments motivated us to revise the standard ChIP-Chip protocol. Analysis of chromosome wide binding of the alternative sigma factor σ32 in Escherichia coli with this new protocol resulted in detection of only a subset of binding sites found in a previous study by Wade and colleagues. We suggested that the remainder of binding sites detected in the previous study are likely to be false positives. In a recent article the Wade group claimed that our conclusion is wrong and that the disputed sites are genuine σ32 binding sites. They further claimed that the non-detection of these sites in our study was due to low data quality.Results/discussionWe respond to the criticism of Wade and colleagues and discuss some general questions of ChIP-based studies. We outline why the quality of our data is sufficient to derive meaningful results. Specific points are: (i) the modifications we introduced into the standard ChIP-Chip protocol do not necessarily result in a low dynamic range, (ii) correlation between ChIP-Chip replicates should not be calculated based on the whole data set as done in transcript analysis, (iii) control experiments are essential for identifying false positives. Suggestions are made how ChIP-based methods could be further optimized and which alternative approaches can be used to strengthen conclusions.ConclusionWe appreciate the ongoing discussion about the ChIP-Chip method and hope that it helps other scientist to analyze and interpret their results. The modifications we introduced into the ChIP-Chip protocol are a first step towards reducing false positive signals but there is certainly potential for further optimization. The discussion about the σ32 binding sites in question highlights the need for alternative approaches and further investigation of appropriate methods for verification.

【 授权许可】

CC BY   
© Schindler and Waldminghaus; licensee BioMed Central Ltd. 2013