# Obtain methylation signatures

The significant CpG loci detected in the previous step actually form a methylation signature that differentiates between LCLs and B cells. We can build and visualize this methylation signature using clustering and a heat map.

* Select the *LCLs\_vs\_Bcells\_CpG\_Islands* spreadsheet in the spreadsheet pane on the left
* Select **Cluster Based on Significant Genes** from the *Visualization* panel of the *Illumina BeadArray Methylation* workflow
* Select **Hierarchical Clustering** for *Specify Method* (Figure 1)

![](https://1384254481-files.gitbook.io/~/files/v0/b/gitbook-x-prod.appspot.com/o/spaces%2FJVEESmJAPppJ3ijFq5aR%2Fuploads%2Fgit-blob-b9098e931cfb6b59ff7770fd004bd3a55e408e70%2Ffigure29.png?alt=media)

Figure 1. Selecting Heirarchical Clustering for clustering method

* Select **OK**
* Verify that **LCLs\_vs\_Bcells\_CpG\_Islands** is selected in the drop-down menu
* Verify that **Standardize** is selected for *Expression normalization* (Figure 2)

![](https://1384254481-files.gitbook.io/~/files/v0/b/gitbook-x-prod.appspot.com/o/spaces%2FJVEESmJAPppJ3ijFq5aR%2Fuploads%2Fgit-blob-51cca550c09cd90d5a561db29be4a29336ccd021%2Fimage2017-12-26%2012_3_34.png?alt=media)

Figure 2. Selecting spreadsheet and normalization method for clustering

* Select **OK**

The heat map will be displayed on the Hierarchical Clustering tab (Figure 3).

![](https://1384254481-files.gitbook.io/~/files/v0/b/gitbook-x-prod.appspot.com/o/spaces%2FJVEESmJAPppJ3ijFq5aR%2Fuploads%2Fgit-blob-9d8094d352ae0b8c0269adb3f5fd45c62411abec%2Fimage2017-12-26%2012_31_33.png?alt=media)

Figure 3. Hierarchical clustering with heat map invoked on a list of significant CpG loci

The experimental groups are rows, while the CpG loci from the *LCLs vs B cells* spreadsheet are columns. Methylation levels are compared between the LCLs and B cells groups. CpG loci with higher methylation are colored red, CpG loci with lower methylation are colored green. LCLs samples are colored orange and B cells samples are colored red in the dendrogram on the the left-hand side of the heat map.

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