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+
+ 
+
+
+raw data produced in Tellini et al., can be downloaded [here](http://134.59.51.17:5000/sharing/keXp2tuto)
+
+:construction: this page is under construction (31/07/2023)
+
+:rocket: A v.2 with several improvements in stability, speed and memory consumption is close to being released.
+
+# intropipeline
+
+[](https://github.com/nicolo-tellini/intropipeline/blob/main/LICENSE)
+[](https://github.com/nicolo-tellini/intropipeline/releases/tag/v.1.0.0)
+[](https://github.com/nicolo-tellini/intropipeline/releases/tag/v.1.0.0)
+[](https://github.com/nicolo-tellini/intropipeline/graphs/commit-activity)
+
+An automated computational framework for detecting *Saccharomyces paradoxus* introgressions in *Saccharomyces cerevisiae* strains from paired-end illumina sequencing.
+
+
+ 
+
+
+## Description
+
+v1. is described in Tellini et al 20xx for detecting *S.par* introgressions in *S.cer* strains.
+
+v2. contains the following implementations and changes:
+- ```minimap2``` replaced ```bwa mem``` almost halving the running time (see [Heng Li 2018, Bioinformatics](https://academic.oup.com/bioinformatics/article/34/18/3094/4994778?login=true)) achieving comparable results;
+
+ sample: ERR3010122
+
+ threads: 2
+
+ Architecture: x86_64
+
+ CPU: Intel(R) Core(TM) i9-9900K CPU @ 3.60GHz
+
+
+ | script | Elapsed Time | Maximum resident set size (GB) |
+ | ------------- | ------------- | ------------- |
+ | bwa mem + samtools (v1) | 6:21 (m:ss) | 1.3 |
+ | minimap2 + samtools (v2) | 3:36 (m:ss) | 1.3 |
+
+- improved the reproducibility of the mapping by implementing the standard samtools workflow according to [samtools' guideline](http://www.htslib.org/workflow/fastq.html)
+- improved the roboustness of the mapping by appending the name of the strain to a checkpoint (cps) file (```./cps/cps.txt```). The strains which names are stored in ```./cps/cps.txt``` will not be mapped again.
+- introduced ```data.table```, ```lapply``` and custom function for large file manipulation for reducing runtime and RAM load.
+ example:
+
+ sample: ERR3010122
+
+ threads: 2
+
+ Architecture: x86_64
+
+ CPU: Intel(R) Core(TM) i9-9900K CPU @ 3.60GHz
+
+ | script | Elapsed Time (s) | Maximum resident set size (GB) |
+ | ------------- | ------------- | ------------- |
+ | parser_marker.r (v1) | 0:17 s | 0.8 |
+ | parser_marker.r (v2) | 0:06 s | 0.5 |
+ | clrs.r (v1) | 0:49 s | 1.9 |
+ | clrs.r (v2) | 0:17 s | 0.7 |
+
+- introduced the variables ```nSamples``` and ```nThreads``` inside ```runner.sh```. The first variable controls the number of samples to run in paralell and the second the per-samples number of threads. ```nSamples``` guarantees a contant number of samples running in parallel; as soon as the count drop of one sample an other will start to run. The definition of these variables affect the scripts ```minimap2.sh``` (which replaces ```bwa.sh```), ```bcftools_markers.sh``` (which replaces ```samtools_marker.sh```) and ```freec.sh```;
+- corrected an error that prevented the detection of the CNVs;
+- Added a new approach for merging markers in blocks:
+
+ In v1 the markers are (1) genotyped, (2) filtered and (3) joined as long as they are consecutive and carry the same information. In v2 this does not change.
+
+ In v2 the markers are (1) ranked, (2) genotyped, (3) filtered, (4) joined as long as they are consecutive in the **ranking** and carry the same information. v1 did not use the ranking.
+ Inevitably, this results in a more fragmented signal but provides a more realistic and faithful representation of the introgression reflecting regions where the genotyping was either discordant or failed.
+ The ranking also represents the strategy that allowed the speedup of ```clrs.r``` (the script that generates the blocks).
+
+
+ 
+
+
+## Download
+
+:octocat: :
+
+```sh
+git clone --recursive https://github.com/nicolo-tellini/intropipeline.git
+```
+
+## Content
+
+:open_file_folder: :
+
+```{bash}
+.
+├── rep
+│ ├── Ann
+│ └── Asm
+├── runner.sh
+├── scr
+└── seq
+
+5 directories 1 file
+```
+
+- ```rep``` : repository with assemblies, annotations and pre-computed marker table,
+- ```runner.sh``` : the script you edit and run,
+- ```scr``` : scripts,
+- ```seq``` : put the FASTQs files here,
+
+### Before starting
+
+``` gzip -d ./rep/mrktab.gz ```
+
+``` gzip -d ./rep/Asm/*gz```
+
+### About the fastqs
+
+Move the FASTQs inside ```./seq/```
+
+Paired-end FASTQs data **must** be gziped and suffixed with **.R1.fastq.gz** and **.R2.fastq.gz**.
+
+### Default
+
+```./scr/bwa.sh``` uses 2 thread for sample (n.samples = 2).
+
+```./scr/samtools_markers.sh``` uses 1 thread for sample (n.samples = 4).
+
+```./scr/gem.sh``` uses 2 threads.
+
+```./scr/freec.sh``` uses 4 threads.
+
+these values can be changed editing the scripts.
+
+### How to run
+
+Edit ```runner.sh``` :page_with_curl:
+
+```{bash}
+#!/bin/bash
+
+#####################
+### user settings ###
+#####################
+
+## S. paradoxus reference assembly
+
+ref2Label="CBS432" ## choose the Spar assembly you think better fit the origin of your samples
+
+## short labels (used to name file)
+
+ref2="EU" ## choose a short name for Spar
+
+# STEP 1
+fastqQC="yes" ## fastqc control (required) ("yes","no" or "-" the last is skip)
+
+# STEP 2
+shortReadMapping="yes" ## ("yes","no")
+
+# STEP 3
+mrkgeno="yes" ## ("yes","no")
+
+# STEP 4
+cnv="yes" ## ("yes","no")
+
+# STEP 5
+intro="yes" ## ("yes","no")
+
+#####################
+### settings' end ###
+#####################
+```
+
+Run ```runner.sh``` :runner:
+
+```{bash}
+nohup bash runner.sh &
+```
+## The result
+
+The results concerning the introgressions are stored in ```./int```
+
+Ex.
+
+An Alpechin strain:
+
+
+ 
+
+
+
+## How to interprer the result
+
+Blue-Red plots provides an overview of potential introgressed DNA across the genome.
+The interpretation of the results is a process that require the integration of different data the pipeline produces.
+
+
+ 
+
+
+:exclamation: Reminder: blocks are defined as consecutive markers besring the same genomic info (Homo S.cer, Homo S.par, Het).
+
+
+
+How are markers distributed inside the *S.par* block?
+
+A couple of possible scenarious:
+
+**Case 1**: abundant markers suporting the block
+
+ 
+
+
+:exclamation: Note: Only a few markers in the figure above are represented in the cartoon;
+
+
+**Case 2**: *not* so abundant markers suporting the block
+
+ 
+
+
+:exclamation: Note: you should *not* exclude the possibility that a large events is supported by a low number of markers as in the example.
+
+The number of markers supporting the blocks, the marker density and the info concerning the genotype are stored in ```int``` and ```int/AllSegments```.
+
+## Dependencies
+
+### Softwares
+
+* [FastQC](https://github.com/s-andrews/FastQC/releases/tag/v0.11.9) v. 0.11.9
+* [bwa](https://github.com/lh3/bwa/releases/tag/v0.7.17) v. 0.7.17-r1198-dirty
+* [samtools](https://github.com/samtools/samtools/releases/tag/1.9) v. 1.9
+* [GEM](https://sourceforge.net/projects/gemlibrary/files/gem-library/Binary%20pre-release%203/) v. 1.315 (beta)
+!! The GEM version used for the analyses is 1.759 (not available anymore).
+* [Control-FREEC](https://github.com/BoevaLab/FREEC/releases/tag/v11.6) v. 11.6; makeGraph.R script was renamed makeplotcnv.R; A copy of all the scripts in [FREEC/scripts/](https://github.com/BoevaLab/FREEC) is in scr. Nevertheless freec has to be installed
+* A copy of [sambamba](https://github.com/biod/sambamba/releases/tag/v0.6.5) v. 0.6.5 is provided with the pipeline (no installation required)
+
+### R libraries
+
+* [data.table](https://rdocumentation.org/packages/data.table/versions/1.14.2) v. 1.14.2
+* [ggplot2](https://github.com/tidyverse/ggplot2/releases/tag/v3.3.5) v. 3.3.5
+* [vcfR](https://github.com/knausb/vcfR/releases/tag/v1.12.0) v. 1.12.0
+* [scales](https://cran.r-project.org/src/contrib/Archive/scales/) v. 1.1.1
+* [rtracklayer](http://www.bioconductor.org/packages/3.11/bioc/html/rtracklayer.html) v. 1.48.0
+* [seqinr](https://cran.r-project.org/src/contrib/Archive/seqinr/) v. 4.2-8
+
+## Find out more
+
+*S.cer* consensus assembly [link method paper]
+
+Marker definition [link method paper]
+
+## Citations
+
+## Release history
+
+* v1.0.0 released on 2023
+
+## TO-DO list
+
+### short-term updates
+- rename columns names ClrS
+
+### long-term updates
+
+- more extensive mapping
+- migration to data.table
+- adopt ranking strategy
+- integration genomics annotations
+- traffic light bash (control over the number of samples and threads)
+- switch CNVs