README.md

# Vavassori_LNP2022_RNAseq_lnps

Vavassori V, Ferrari S, Beretta S, et al.
**Lipid nanoparticles improve ex vivo gene editing of human hematopoietic cells.**
2022.

Whole transcriptomic analysis 24 hours after treatment of CD4+ T cells from 3 healthy donors and of HSPC (form 3 replicates) for CD40LG editing treated with LNPs. As controls, cells untreated (UT), mock electroporated or treated with LNPs formulated without RNA (Empty LNPs) were employed.

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### Analyses ###

Stanrdard RNA-seq pipeline on 36 samples: 18 samples of T cells and 18 samples of HSPCs.<br>
In details, for both cell types (i.e., T and HSPC) DGE analysis on samples was performed with DESeq2, comparing dfferent groups (adding donor covariate for T cells):
```
results(DESeq.ds, pAdjustMethod = "BH", independentFiltering = TRUE, contrast = c("groups", Group1, Group2), alpha = 0.05)
```
Enrichment on (positive or negative) differentially expressed genes (FDR < 0.05) and GSEA on pre-ranked gene lists (ordered by log2 Fold Change values) agasint the MSigDB Hallmark database were performed with ClusterProfiler:
```
enricher(PosDEGs, universe = GeneUniverse, pvalueCutoff = 0.05, TERM2GENE = HallMarkDB)

GSEA(FullGenesOrderLogFC, TERM2GENE = HallMarkDB, nPerm = 10000, pvalueCutoff = 0.05)
```

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### Directories and Files ###

- _paper_plots.R_: script used to produce the plots of the paper;
- _paper_data_: folder with data used to produce the plots of the paper;
- _GEO_RNAseq_lnp.xlsx_: GEO template file;
- _config.yaml_: config files of the RNA-seq analysis pipeline.