Spring 2023 Next Generation Sequencing Workshops

These workshops were supported in part by the University of Pittsburgh seed project titled "University of Pittsburgh Computational Genomics Training Program".

High throughput sequencing has brought abundant sequence data along with a wealth of new “-omics” protocols, and this explosion of data can be as bewildering as it is exciting. Our multi-day hands-on workshops give researchers the research, open-sourced tools to plan and execute successful bioinformatics and genomics experiments. These workshops, taught by experienced Bioinformatics core faculty, cover both the theoretical and practical aspects of a wide range of NGS data, using the HTC cluster.

These workshop have hands-on components that require the following requirements be set up before a workshop begins.

  1. Participants should have an account on the HTC cluster, which is the cluster we will use for demonstration purposes. (page 1 of this documentation)
  2. This workshop also requires that participants either be on a Pitt network (hard-line) or behind a VPN. (page 2 of this documentation)
  3. You can submit jobs, i.e., your group’s account has not expired, and your group’s service units (CPU-hours) have not been exhausted entirely (page 4 of this documentation)

As a general rule, we offer no troubleshooting for technical setup issues at the workshops themselves! Therefore, be aware that if you do not set up the workshop's technical prerequisites well in advance, you may not be able to participate fully in its hands-on activities.

You can find the titles of past NGS workshops and links to their recordings. For each workshop, there may also be slides used in the workshop and other additional relevant resources.

A familiarity with Linux and the Bash Shell is vital for these workshops. Submitting, monitoring, and managing jobs on the HTC cluster largely involves command-line operations. We do not routinely teach beginning Linux classes. If you are new to Linux environments, we highly recommend that you work your way through one of the past recordings (Introduction to Linux for NGS, Spring or fall 2021 workshop).

Register for all workshops: Spring 2023 NGS workshops

Next Generation Sequencing Techniques

Tuesday, Jan. 24, 2023
1:00 pm - 2:30pm

This workshop will cover the basis of Next-Gen Sequencing Library Preparation for Illumina Sequencers. Different Library Preparation Techniques (DNA-seq, ChIP-seq, RNA-seq, Methyl-seq and Spatial Transcriptomics) are explained. Quality Control steps of the starting input material and final libraries are also explained. This workshop will also discuss considerations for experimental design and the end goals of analysis prior to sequencing. Basics of sequencing and cost estimates will be discussed in the experimental design process. Presented by Amanda Poholek

T cell receptor (TCR) data analysis

Tuesday, Jan. 31, 2023
1:00pm - 4:00pm

In this workshop, we will focus on characterizing tumor-infiltrating T cell receptor (TCR) repertoire from bulk RNA-sequencing data. The workflow will cover the implementation of computational algorithms to extract TCR hypervariable complementarity determining region 3 (CDR3) followed by descriptive statistics for TCR repertoires, shared clonotype analysis and repertoire comparison, repertoire diversity and gene usage analysis, and visualization. Presented by Dhivyaa Rajasundaram.

ATACseq data analysis

Tuesday, Feb. 7, 2023
1:00pm - 4:00pm

This workshop will present the ATACseq (Assay for Transposase-Accessible Chromatin with high-throughput sequencing) method. The focus of the workshop will be on running ATAC-seq pipelines, from raw fastq files, to quality control, alignment to reference genome, peak calling and differential analysis. nf-core ATACSeq pipelne will also be introduced. Presented by Fangping Mu

Methylation data analysis

Tuesday, Feb. 14, 2023
1:00pm - 4:00pm

In this workshop, we will focus on an introduction to DNA methylation and whole genome bisulfite sequencing. The workflow will cover data formats, quality control, read mapping, overview statistics, segmentation, windowing, and smoothing. We will also focus on the identification of differentially methylated regions, and visualization of DNA methylation data using R. Presented by Dhivyaa Rajasundaram.

RNAfusion data analysis

Tuesday, Feb. 28, 2023
1:00pm - 4:00pm

This workshop will focus on the detection of fusion transcripts from sequencing data. The first half of the workshop will be a theoretical introduction. We will share some basic concepts of the fusion transcript and their clinical applications. In addition, we will introduce some bioinformatics pipelines to detect fusion transcripts from next-generation short-read and long-read sequencing data. The second half of the workshop will be a practical section. We will hand on real RNA-seq data, run the pipeline and check the results for fusion detection. We will introduce both script-based fusion detection tools, as well as their nf-core pipelines for user-friendly fusion detection and visualization. Presented by Silvia Liu

Nanostring spatial transcriptomics data analysis

Thursday, March 16, 2023
1:00pm - 4:00pm

This workshop will provide basic principle of Nanostring spatial transcriptomics and pipelines for data analysis. In the first part of the workshop, we will introduce this cutting-edge technology, briefly review several applications in research studies, check the data format and illustrate bioinformatics data analysis pipelines for pre-processing and downstream analysis. In the second part, we will apply Seurat package to work on real data. Presented by Silvia Liu

10X Genomics single cell techniques     

Monday, March 27, 2023
1:00pm – 2:30pm

This workshop will illustrate the technological basis of 10x Genomics single cell analysis. Several chemistries for applications beyond single cell RNA-Seq and the sequencing read architecture of each application will be discussed. Presented by Richard Duerr

Single-cell Analysis with Seurat  

Tuesday, March 28, 2023
1:00pm – 4:00pm

This workshop will briefly review the cellranger pipelines to process raw reads into expression values. The hands-on training will include reading the count data in R, quality control, normalization, dimensionality reduction, cell clustering, and finding marker genes. The Seurat pipeline will be covered. We will also focus on trajectory analysis with Monocle3. Presented by Dhivyaa Rajasundaram

Single-Cell Analysis with Bioconductor   

Tuesday, April 4, 2023
1:00pm – 4:00pm

This workshop will teach how to perform scRNAseq analysis using Bioconductor. The training will include many aspects of analysis such as normalization, cell type annotation of clusters, and pseudo-bulk differential gene expression. Similarities and differences to the Seurat pipeline will be mentioned. Presented by Paul Cantalupo (Genomics Analysis Core)

Annotating single-cell transcriptomics

Thursday, April 6, 2023
1:00pm – 4:00pm

Single-cell transcriptomics can quantitatively measure and identify cell types and dynamics based on gene expression profiles. Accurate annotation of cells is the foremost step for downstream analyses. In this workshop, we will focus on how to annotate different cell populations in human biospecimens using automatic or manual approaches, and detect cellular states for biological interpretation. We will also discuss the recommendations, challenges, and best practices for the selection of reference databases, probability models, and relevant computational tools. Presented by Riyue Bao

Advanced Single-Cell Analysis 

Tuesday, April 11, 2023
1:00pm – 4:00pm

This advanced workshop will focus on the analysis of multi-omics data and its integration (e.g. CITE-seq, multiome, and DOGMA-seq). The hands-on training will include background introduction and a step-by-step R pipeline with real applications from immunology. Presented by Wei Chen

Deep learning for scRNASeq analysis

Tuesday, April 25, 2023
1:00pm – 4:00pm

This workshop will review the deep learning algorithms (variational autoencoder, autoencoder, generative adversarial network and supervised DL models ) and their applicability in the single cell RNA-seq processing pipeline. The hands-on training will provide examples to run deep learning RNASeq algorithms.. Presented by Yufei Huang