Bioinformatics- Promoter/H3K27me3 mark crosses in excel
GALAXY Procedure
- Download .bed data from lab's backfile and upload to GALAXY (usegalaxy.org). Toolbar left > Get Data > Upload File > Select .bed file > File Format: Interval, Genome: hg18 > Execute.
- Upload refseq promoters (hg18) in same fashion as .bed data if stored on hard drive.
- Intersect promoter with crosses: Toolbar left > Operate on Genomic Intervals > Intersect > Return: Overlapping Intervals, of: methyl-histone data, that intersect: promoters, for at least: 1 bp > Execute.
- Once new data analysis is run, regions should display under title for analysis. Enter all crosses data into excel spreadsheet for analysis.
- Intersect new cross data with all other crosses made. This will give regions that both cell experiments contain. For example, a cross for cell experiments A1 and B1 will contain all regions that have histone methylation in both data.
Excel Procedure
- Create a table consisting of all cell experiments from alpha cells to exocrine cells on both the x and y axis (Figure 1).
- In each cross (e.g. A1 cross B1) mark down the number of regions similar. Data will come from previous GALAXY crosses.
- Heat map analysis using conditional formatting on Excel. Darker numbers = near maximum, lighter numbers = near minimum.
- Crosses between H3K27me3 data (Bramswig et al. 13)and reference promoters were done in Excel. They show number of genes hypothetically controlled by the histone methylation mark.
- Crosses revealed around 4950 promoters were controlled in alpha cells. Heat map analysis indicates that alpha cells are more heavily affected by H3K27me3 marks than beta cells. Crosses between alpha/beta cell histone methylation (i.e. A1-3 cross B1-3 on the table) show there are around 1000 genes that are associated with the methyl-histone mark in alpha cells but not in bet cells.
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Cell Fate Switch by Synthetic Transcription Factors
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