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Old 05-09-2013, 04:37 AM   #1
Location: haifa israel

Join Date: Jun 2011
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Default Cluster analysis of RNASeq data

I have a data set made with 6 developmental time points during development, on a newly assembled transcriptome. We used BLAST2GO to annotate the trasncriptome. In looking for interesting genes or patterns to follow up on, we wanted to perform clustering. Since this is my first move into clustering, does anyone have recommendations? I have no idea how to choose between hierarchial, kmeans, etc (and if kmeans - which k do I choose?)
Any help (even pointing me in the direction of a good intro paper) would be appreciated!
Also- what kinds of normalization do I need to do to preprocess the different stages? I feel that the FPKMs are a bit higher than average in one stage and a bit lower than average in another and have no idea how to deal with this.
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Old 05-09-2013, 05:37 AM   #2
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There are pros and cons to each approach. Hierarchical will give you a cluster of clusters. This could be an advantage or completely useless. Hierarchical clustering is also prone to outliers if I remember right. An outlier would form its own cluster entirely. On the other hand k-means requires you to define the number of clusters. This can be done by using the Gap statistic of Tibshirani.
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Old 06-13-2013, 07:31 AM   #3
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Hi, Hierachical clustering is a nice way of representing the samples difference and to look at the relationship between the samples in the initial stages of how raw data look like. Although the way the clusters are formed in tree corresponds to how we calculate the distance measure(Single/Wards/Complete/Average) and the type of method (Euclidean/Pearson)used to calculate the distances between two data points . I have 16 RNA-seq samples, tried to perform hierarchical clustering on dataset, by using Euclidean distance measure and Wards methods, the tree genertated was different if i use Single-linkage method. Although by Single linkage method the tree is making sense in terms of biology, but I am not completely clear of the point which will be the optimal method to consider when we look at the RNA-seq data having count values??

Any suggestions please??
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Old 06-13-2013, 09:41 PM   #4
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Default non-negative matrix factorization (NMF)

non-negative matrix factorization (NMF) also allows unsupervised clustering, defining also the number of clusters which represents the data best using the cophenetic correlation index. in my hands this method produces more robust clusters compared to hierarchical clustering...

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