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CALENDAR

June 12–13, 2020

Click here to download the schedule in PDF. The current schedule may suffer small alterations as we get closer to the start of the course. Please be sure to check it again later.

Calendar: Admissões

DAY 1

June 12, 2020

ARRIVAL AND INSTRUCTIONS

10:00 am

  • Introduce instructors and their background

  • Have students introduce themselves

SETUP/TROUBLESHOOT USER SYSTEMS

10:15 am

  • Make sure participants have Oracle VM VirtualBox and the course imaging running on their personal computers

  • Show the location of directories containing course materials (references, list of exercises, and examples)

EXPERIMENTAL DESIGN

11:15 am

  • Choosing among high-throughput RNA sequencing technologies

  • Strategies for RNA extraction, quantification, and storage

  • Considerations for sequencing library choice and preparation

  • Calculating expected coverage to optimize costs

  • Multiplexing strategies to optimize sequencing from multiple samples

  • Cost estimation

  • Exercise list #1 (coverage calculation and cost estimation)

LUNCH

12:00 am

  • On your own

TOOLS FOR RNA-SEQ DATA ANALYSIS AND QUALITY CONTROL

1:00 pm

  • Summary of the most used open-source/ free software for transcriptome assembly RNA-Seq analysis

  • Why and how to perform quality control (e.g., adapter trimming and filtering by size)

  • Detection and removal of technical sequences

  • Accessing the quality of each independent biological replication

  • Exercise list #2 (adapter trimming)

TRANSCRIPTOME ASSEMBLY

1:45 am

  • The bases of reference and de novo assembly

  • RNA-Seq de novo sequence assembly using Trinity

  • Estimating assembly completion using BUSCO

  • Exercise list #3 (sequence assembly)

NORMALIZATION AND DIFFERENTIAL EXPRESSION ANALYSIS

2:30 pm

  • Trimmed Mean of M-values (TMM) normalization method (edgeR)

  • Using RSEM to estimate transcript abundance

  • Identifying differentially expressed (DE) transcripts using edgeR

  • RPKM vs. FPKM

  • Exercise list #3 (estimating the abundance of the individual transcripts)

COFFEE BREAK

3:15 pm

  • On your own

SPLICING ANALYSIS AND MULTIPLE TESTING

3:30 pm

  • The BLAST like alignment tool (BLAT)

  • Spliced aligners (Tophat, GSNAP, SOAPsplice, MapSplice, TrueSite, star)

  • Aligners that can "clip" (Bowtie2's local mode, bwa-mem)

  • Exercise list #5 (an example BLAT analysis)

BIOLOGICAL INTERPRETATION

4:15 pm

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WRAP UP

5:00 pm

  • Gather preliminary feedback from participants and resolve any issues with the course image or the first five lists of exercises.

ADJOURN

5:15 pm

Schedule Item Body

Calendar: Agenda

DAY 2

June 13, 2020

8:30 am

INSTRUCTIONS FOR DAY TWO

  • Introduce speakers of day two and their backgrounds

  • Check that all course participants have a working image on their VMs

  • Show the location of directories containing course materials for this second day of the short course (references, list of exercises, and examples)

9:15 am

INTRODUCTION TO GENE ANNOTATION

  • Difference between gene prediction and annotation

  • The importance of gene annotation of comparative analysis

  • How to read annotation data from genomic sequences published in GenBank (.gb) format

  • Exercise list #6 (reading gene annotations)

10:00 am

COFFEE BREAK

  • On your own

10:30 am

AB INITIO GENE ANNOTATION

  • Definition of ab initio (intrinsic) annotation

  • Pros and cons (when to use it, why, and what to expect)

  • Sites we might want to predict (translation start, splice donor site, splice acceptor site, translation stop)

  • Hidden Markov Models (HMMs)

  • Exercise list #7 (examples of gene prediction using GENSCAN and Augustus)

11:15 am

HOMOLOGY-BASED GENE ANNOTATION

  • Definition of homology-based (extrinsic) annotation

  • Pros and cons (when to use it, why, and what to expect)

  • How to calculate and interpret the E-value and the bit-score

  • Exercise list #8 (examples of gene prediction using ORFfinder, BLASTN, and TBLASTN; comparison with results from exercise list #2)

12:00 am

LUNCH

  • On your own

1:00 pm

VISUALIZING GENE ANNOTATION

  • Examples of open-source or free tools to visualize gene annotation data

  • Exercise list #9 (Generic Genome Browser - GBrowser and Integrative Genomics Viewer - IGV)

1:45 pm

SHARING GENE ANNOTATION

  • Different formats for sharing gene annotation, with focus on the Generic Feature Format version 3 (GFF3)

  • Exercise list #10 (filtering, concatenating, and extracting information from GFF3 files using simple pre-written Python scripts)

2:30 pm

COFFEE BREAK

  • On your own

2:45 pm

MANAGING DATA FROM PUBLIC DATABASES

  • List of main public resources

  • How to leverage on EchinoDB for your research

  • The importance of using the correct gene names

  • Exercise list #11 (comparing your gene annotation with information available in EchinoDB to identify the gene of interest and compare it with orthologous sequences)

3:30 pm

WRAP UP

  • Gather feedback from participants on how the course will be of use to their work and general course feedback

3:45 pm

ADJOURN

Schedule Item Body

Calendar: Agenda
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