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# ikra v1.2.0 -RNAseq pipeline centered on Salmon-<imgsrc="img/ikra.png"width="20%"align="right" />
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# ikra v1.2.1 -RNAseq pipeline centered on Salmon-<imgsrc="img/ikra.png"width="20%"align="right" />
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A gene expression table (gene × sample) is automatically created from the experiment matrix. The output can be used as an input of [idep](http://bioinformatics.sdstate.edu/idep/). Ikra is an RNAseq pipeline centered on [salmon](https://combine-lab.github.io/salmon/).
-**test option** limits the number of reads to 100,000 in each sample.
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-**udocker mode** is for server environments that can only use User privileges. For more information [https://github.com/indigo-dc/udocker](https://github.com/indigo-dc/udocker).
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-**without-docker mode** works with all tools installed. Not recommended.
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-**protein-coding mode** restricts genes to protein coding genes only.
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-**threads**
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-**output** is `output.tsv` by default.
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**experiment matrix** should be separated by commas (csv format).
-Denote names by connecting conditions and replicates with underscores. See [idep's Naming convention](https://idepsite.wordpress.com/data-format/) in detail.
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-The first three columns are required.
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-If you want to use your own fastq file, add `--fastq` option. Ikra supports only `.fq`, `.fq.gz`, `.fastq` and `fastq.gz`.
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- fastq file specifies path excluding `fastq.gz` or `_1.fastq.gz` and `_2.fastq.gz`. For example, `hoge/SRR5385247.fastq.gz` is described as `hoge/SRR5385247`.
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-If suffix is not `_1.fastq.gz` or `_2.fastq.gz`, add -s1 and -s2 options.
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-It is impossible for docker to specify a hierarchy above the execution directory, such as `../fq/**.fastq.gz`, but it can be avoided by pasting a symbolic link.
- Ion S5用: SEしか無い。trimmomaticではなくfastx-toolsを使う。adapterはNoneを入れておく。(test : [DRP003376](https://trace.ncbi.nlm.nih.gov/Traces/sra/?study=DRP003376))
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### Output
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- output.tsv(scaledTPM)
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- multiqc_report.html : including fastQC reports and mapping rate of salmon(mapping rate for transcript)
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**output sample**
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|| Treg_LN_1 | Treg_LN_2 |
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| ---- | ---- | - |
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| 0610005C13Rik | 0 | 0 |
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| 0610006L08Rik | 0 | 1 |
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| 0610009B22Rik | 4 | 10 |
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| ... |||
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- multiqc_report.html
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salmonのマッピング率（トランスクリプトに対するマッピング率）
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### Various Specifications
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### 各種仕様
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- output is **scaledTPM** (see. [Soneson, C., Love, M. I. & Robinson, M. D. Differential analyses for RNA-seq: transcript-level estimates improve gene-level inferences. F1000Research 4, 1521 (2015).](https://f1000research.com/articles/4-1521/v2))。
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- About GCbias `—-gcbias` is added on salmon. You can refer to https://mikelove.wordpress.com/2016/09/26/rna-seq-fragment-sequence-bias/ about the influence on RNAseq by GCbias.
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- ValidateMappings option was adopted. (You can’t use it while using alignment-base mode.) Please see https://combine-lab.github.io/salmon/faq/ for further details.
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- outputは**scaledTPM** (see. [Soneson, C., Love, M. I. & Robinson, M. D. Differential analyses for RNA-seq: transcript-level estimates improve gene-level inferences. F1000Research 4, 1521 (2015).](https://f1000research.com/articles/4-1521/v2))。
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- GCbiasについて、salmonで`--gcBias`を追加した。GCbiasのRNAseqにおける影響に関しては[Mike Love's blog :
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RNA-seq fragment sequence bias](https://mikelove.wordpress.com/2016/09/26/rna-seq-fragment-sequence-bias/)。
A serious bug was reported in the `tximport_R.R` and fixed. In the older version, Salmon's output and multiqc reports were correct and sometimes `output.tsv` were disturbed. Please update Ikra to the latest version. If you are using the old version(<1.1.1), please update and re-run ikra. We apologize for the inconvenience.
All you need is `git clone` ikra, and install docker or udocker(v1.1.3). No need for installing plenty of softwares! If you don’t want to use docker (or udocker), you must install all softwares by yourself and use `—-without-docker` option.
[Dr.Ota(DBCLS)](https://github.com/inutano) solved the problem that salmon doesn’t work on Mac. The cause of the problem is that Docker is allocated only 2GB by default on Mac. Therefore,like this picture, the problem will be solved by allocating large amount of memories to Docker, such as 8GB, and doing Apply & Restart.
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