From cdb5acb53a1d6c27a8c31811827074e71aa9c1a6 Mon Sep 17 00:00:00 2001
From: Pariksheet Nanda <pariksheet.nanda@uconn.edu>
Date: Fri, 17 Mar 2017 00:34:41 -0400
Subject: [PATCH] DOC: Add README

---
 README.md | 118 ++++++++++++++++++++++++++++++++++++++++++++++++++++++
 1 file changed, 118 insertions(+)
 create mode 100644 README.md

diff --git a/README.md b/README.md
new file mode 100644
index 0000000..a43e055
--- /dev/null
+++ b/README.md
@@ -0,0 +1,118 @@
+Centromere Transcription Factor RNAi screen
+===========================================
+
+Search for genes that affect centromere establishment.
+
+Experiment background
+---------------------
+
+Centromeres are essential for cell division.
+They recruit the proteins to which the spindle fibers attach,
+and the spindle fibers segregate the sister chromatids.
+
+Centromeres require a unique sub-unit protein
+in their histone octamer called CENP-A.
+After each cell division, the CENP-A histone proteins
+need to replace the H2A-H2B dimer to reform the centromere.
+This process is thought to require transcription.
+
+
+Experiment summary
+------------------
+
+To study centromere formation
+one needs to seed for the protein complex
+that will incorporate CENP-A into histones,
+replacing the original H2A-H2B histone sub-units.
+The "seed" to trigger the events of this histone replacement
+is a Drosophila specific protein called CAL1.
+CAL1 is brought to the ectopic centromere site
+using the LacO-LacI tethering system:
+namely, several (32) LacO repeats
+are inserted into a region of chromosome 3L
+and LacI-CAL1 will be recruited to that region
+due to the LacO-LacI affinity.
+About 25% of the time, this system creates a new centromere.
+
+The experiment searches for proteins involved
+with centromere formation by knocking down known nuclear proteins,
+which include transcription factors.
+When ectopic formation dips significantly below the typical 25%,
+we may have found a gene involved with centromere formation.
+
+The data collected are fluorescent images of multi-well plates,
+where each well corresponds to a protein being knocked down.
+
+  - A red fluorescent protein labels the ectopic centromere location
+    (of the LacO repeats).
+  - A green fluorescent protein labels CENP-A.
+  - DAPI blue fluorescence labels nuclear DNA.
+
+Usage
+-----
+
+Run the `Makefile` in this directory to generate all results.
+
+``` sh
+make
+```
+
+To tune CellProfiler's image processing,
+it is helpful to save results in a separate directory.
+One can clone this repository using a different directory name,
+and link to the processed image set of the original:
+
+``` sh
+cd ..
+git clone git@github.uconn.edu:MelloneLab/RNAi_plate_analysis_all.git rnai-screen-tf_20170314
+cd rnai-screen-tf_20170314
+rm -rf z_projection
+ln -s ../../rnai-screen-tf/results/z_projection z_projection
+```
+
+A nice feature of Makefiles is the ability to overwrite any number of variables
+by specifying it on the command-line in the general form `variable=value`.
+For example, to use cellprofiler installed to your personal directory
+by `pip install --user ...`,
+you may specify the path to cellprofiler as:
+
+``` sh
+make CELLPROFILER=~/.local/bin/cellprofiler
+```
+
+Data processing
+---------------
+
+The raw input data consists of:
+
+  1. Images of 5 plates, with 10 sites per well
+	 and 3 z-slices per site ("April_16_2016.tar.xz").
+  2. A spreadsheet mapping the proteins to the wells
+     ("DRSC_TF_Library_Distribution.xls").
+
+Below is the file listing of the "data" directory using `tree`:
+
+```
+data
+├── April_14_2016 [11 entries exceeds filelimit, not opening dir]
+├── April_14_2016.tar.xz
+└── DRSC_TF_Library_Distribution.xls
+```
+
+The images are 19 GB in the xz compressed archive,
+therefore it is download from FIXME_INSERT_DOI
+to the data directory.
+
+Broadly speaking this data is processed as follows:
+
+  1. CellProfiler saves image statistics to a database.
+  2. R scripts to save high confidence wells and generate plots.
+
+CellProfiler 2 requires 2D image inputs,
+therefore a Python script creates the z-projections.
+
+CellProfiler segments the ectopic and CENP-A centromeres and
+saves the statistics into an sqlite database.
+
+The R-scripts read this CellProfiler generated database for their calculations
+and plots.