Source: UNIV OF MARYLAND submitted to NRP
HIGH-QUALITY REFERENCE ASSEMBLY AND ANNOTATION OF THE RAINBOW TROUT GENOME
Sponsoring Institution
National Institute of Food and Agriculture
Project Status
COMPLETE
Funding Source
AFRI COMPETITIVE GRANT
Reporting Frequency
Annual
Accession No.
1021456
Grant No.
2020-67015-30770
Cumulative Award Amt.
$500,000.00
Proposal No.
2018-06539
Multistate No.
(N/A)
Project Start Date
Apr 1, 2020
Project End Date
Dec 31, 2024
Grant Year
2020
Program Code
[A1201]- Animal Health and Production and Animal Products: Animal Breeding, Genetics, and Genomics
Recipient Organization
UNIV OF MARYLAND
(N/A)
COLLEGE PARK,MD 20742
Performing Department
Animal and Avian Sciences
Non Technical Summary
With sustained high rates of human population growth and depletion of natural fisheries resources, the US seafood imports are expected to continue to increase sharply. Aquaculture is the world's fastest-growing agri-food business. Rainbow trout is the most cultivated cold-water fish in the US. The US aquaculture industry needs applications of genomics in breeding programs to define genetic and phenotypic parameters that control complex traits such as disease resistance and feed efficiency. The FAASG international consortium "Functional Analysis of All Salmonid Genomes" has been established to study the functional genomic basis of phenotypic variation in all salmonids including rainbow trout (https://www.faasg.org/). A trout reference genome sequence is now available (GenBank Accession GCA_002163495). However, major improvements in genome reference and annotation can improve the accuracy and efficiency of the genomic applications such as genome-wide association studies, identification of causative variations, marker-assisted selection and genomic selection in rainbow trout. Therefore, the objectives of our current USDA-supported project are: 1) Closing the Swanson reference genome assembly by utilizing the long-read PacBio sequencing platform in combination with the BioNano whole-genome optical map. 2) Annotate the reference genome for the coding and non-coding transcript isoforms and alternative splicing, by full-length single-molecule sequencing. 3) Annotate the genome for chromatin histone modifications and chromatin accessibility by integrating data from RNA-seq, DNAse-seq, and ChIP-seq across a wide range of rainbow trout tissues. The improved assembly and annotation of the reference genome will accelerate the genetic selection efforts, particularly through GWAS and genomic selections, for improving important production traits in rainbow trout.
Animal Health Component
40%
Research Effort Categories
Basic
10%
Applied
40%
Developmental
50%
Classification

Knowledge Area (KA)Subject of Investigation (SOI)Field of Science (FOS)Percent
30408101080100%
Knowledge Area
304 - Animal Genome;

Subject Of Investigation
0810 - Finfish;

Field Of Science
1080 - Genetics;
Goals / Objectives
The purpose of this project is to provide a high-quality assembly and well-annotated reference genome of the rainbow trout. The new genome reference will accelerate the genetic selection efforts, particularly through GWAS and genomic selections, for improving important production traits in rainbow trout.
Project Methods
Closing the reference genome assembly by utilizing the long-read PacBio sequencing platform in combination with the BioNano whole-genome optical mapping, Dovetail HiRise map of chromatin interactions and genetic mapping.Generate ~120X sequence coverage of the genome using the PacBio long-read technology to develop a new RBT genome assembly with much better contig size.Scaffolding of the contigs from PacBio and from our most recent Illumina-based assembly (reference genome Accession GCA_002163495) with optical mapping using the Irys optical mapping technology (BioNano Genomics) and the previously generated Dovetail HiRise map of chromatin interactions. The PacBio and BioNano technologies will lead to assembling larger scaffolds and better chromosome sequences (anchoring and ordering the majority of the sequences into chromosomes with high accuracy).Use the previously generated linkage map, composed of ~46K SNPs on the 29 chromosomes (collected from 5,716 fish and 146 full-sib pedigreed families), to aid in anchoring of the de-novo assembly scaffolds to chromosomes.Annotate the reference genome for the coding and non-coding transcript isoforms and alternative splicing, by full-length single-molecule Iso-seq sequencing of the transcriptome across a wide range of vital tissues, developmental stages, and genetic lines.Annotate the genome for chromatin histone modifications, chromatin accessibility, by integrating data from RNA-seq, ATAC-seq, and ChIP-seq across various tissues of the rainbow trout.

Progress 04/01/20 to 12/31/24

Outputs
Target Audience:Aquaculture producers, breeders, and scientific community. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?The project provided opportunities to train 4graduate students, a postdoctoral fellow, and several undergraduates. How have the results been disseminated to communities of interest?The results have been published, made publicly available, and presented to stakeholders at scientific conferences. What do you plan to do during the next reporting period to accomplish the goals? Nothing Reported

Impacts
What was accomplished under these goals? The project successfully achieved its objectives: A new chromosome-level genome assembly with gene annotation for the Swanson homozygous line of rainbow trout has been published and is publicly available. A comprehensive catalog of the rainbow trout epigenome, including annotation tracks, has been published and made publicly accessible. These annotation tracks provide key gene regulatory elements, including chromatin histone modifications, chromatin accessibility, and DNA methylation. This regulatory element catalog offers insights into epigenome dynamics and its correlation with gene expression by integrating chromatin immunoprecipitation sequencing, ATAC sequencing, Methyl Mini-seq, and RNA sequencing data. The epigenomic analysis identified potential causal variants and transcription factors influencing complex domestication traits. Additionally, it sheds light on the epigenome's role in gene evolution and the retention of duplicate genes--100 million years after whole-genome duplication in rainbow trout and during re-diploidization. The newly developed epigenome annotation maps, among the first in fish, are expected to enhance the accuracy and efficiency of genomic studies. They will support applications such as genome-wide association studies, causative variant identification, genomic selection in rainbow trout, and fish comparative genomics.

Publications

  • Type: Peer Reviewed Journal Articles Status: Published Year Published: 2024 Citation: Mohamed Salem, Rafet Al-Tobasei, Ali Ali, Liqi An, Ying Wang, Xuechen Bai, Ye Bi, Huaijun Zhou, Functional annotation of regulatory elements in rainbow trout uncovers roles of the epigenome in genetic selection and genome evolution, GigaScience, Volume 13, 2024, giae092, https://doi.org/10.1093/gigascience/giae092
  • Type: Peer Reviewed Journal Articles Status: Published Year Published: 2025 Citation: Ali, A., Gao, G., Al-Tobasei, R. et al. Chromosome level genome assembly and annotation of the Swanson rainbow trout homozygous line. Sci Data 12, 345 (2025). https://doi.org/10.1038/s41597-025-04693-7
  • Type: Conference Papers and Presentations Status: Published Year Published: 2024 Citation: Salem M. Chromatin State and DNA Methylation Profiling of the Rainbow Trout Genome Reveal Regulatory Elements Involved in Controlling Gene Expression and Gene Evolution and epiQTL Signatures of Domestication. PAG- 31 Plant & Animal Genomes Conference (January 12-17, 2024), San Diego, California
  • Type: Conference Papers and Presentations Status: Published Year Published: 2025 Citation: Ahmed, R , Raymo, G , Ali A , Al-Tobasei R, Leeds T, Salem M. Multi-omics analysis identifies candidate genetic and microbial markers and their potential interaction to influence fillet yield and quality traits in rainbow trout. Aquaculture America (March. 6-10, 2025), New Orleans, LA
  • Type: Conference Papers and Presentations Status: Published Year Published: 2025 Citation: Ahmed R , Raymo G , Ali A , Al-Tobasei R, Leeds T, Salem M. Multi-Omics Analysis Reveals Genetic and Microbial Biomarkers and Their Association with Growth Traits in Rainbow Trout. PAG- 32 Plant & Animal Genomes Conference (January 10-15, 2025), San Diego, California.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2025 Citation: Ali Ali , Salem M. Tracing 100 Million Years of Gene Evolution Following Whole Genome Duplication in Rainbow Trout. PAG- 32 Plant & Animal Genomes Conference (January 10-15, 2025), San Diego, California
  • Type: Conference Papers and Presentations Status: Published Year Published: 2025 Citation: Ali Ali , Salem M. New Rainbow Trout Genome Assembly Allows Tracing 100 Million Years of Gene Evolution Following Whole Genome Duplication, the 5th International Conference on Integrative Salmonid Biology (March 11-14, 2024, Seattle, WA)
  • Type: Conference Papers and Presentations Status: Published Year Published: 2024 Citation: Ahmed R O , Ali A, Leeds T, Salem M. Gene Expression Signature, Genetic Polymorphism, And Microbiome Biomarkers Associated With Fillet Pink Color In Rainbow Trout. Aquaculture America (Feb. 18-21, 2024), San Antonio, TX


Progress 04/01/23 to 03/31/24

Outputs
Target Audience:Aquaculture producers, breeders, and scientific community. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?This project provided opportunities to train 1 postdoc, 3 graduate students, and 10 undergraduate students. How have the results been disseminated to communities of interest?Conference presentations andpublications. What do you plan to do during the next reporting period to accomplish the goals?3 major journal papers are ready for submission.

Impacts
What was accomplished under these goals? Three genome assemblies are now available for rainbow trout: NCBI GCA_013265735.3, GCA_025558465.1, and GCA_029834435.1.

Publications

  • Type: Journal Articles Status: Published Year Published: 2023 Citation: Ahmed R. O., Ali A, Leeds T, Salem, M (2023). RNA-Seq Analysis Of The Pyloric Caecum, Liver, And Muscle Reveals Molecular Mechanisms Regulating Fillet Color In Rainbow Trout. BMC Genomics 24, 579
  • Type: Conference Papers and Presentations Status: Accepted Year Published: 2023 Citation: Salem M. An Update on Improving Genome Assembly and Functional Annotation for Rainbow Trout. PAG- 30, Plant & Animal Genomes Conference; (January 13-18, 2023), San Diego, California. https://plan.core-apps.com/pag_2023/abstract/ef218abe-90e0-4afc-8fce-e56385809487.
  • Type: Conference Papers and Presentations Status: Accepted Year Published: 2023 Citation: Salem M. Status of the Assembly and Functional Annotation of Rainbow Trout Genome. Aquaculture America (Feb. 23-28, 2023), New Orleans, LA.


Progress 04/01/22 to 03/31/23

Outputs
Target Audience:Aquaculture and fishscientific community. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?The project helped in training 2 postdocs, 3 graduates, and several undergraduate students. How have the results been disseminated to communities of interest?Results were published in scientific journals and presented in scientific meetings. What do you plan to do during the next reporting period to accomplish the goals?An NCE is necessary to complete the high-throughput integration analyses to deliver a high-quality chromosome-level genome reference with high-quality annotation, including identifying protein-coding and noncoding regions and chromatin accessibility. ATAC-Seq data and Chip-Seq data from six tissues were generated but need to be integrated into the genome to annotate chromatin accessibility for gene expression. A publication detailing project outcomes is in preparation.

Impacts
What was accomplished under these goals? We have finished obtaining DNA and RNA sequences to fulfill most of the following objectives of this project: 1) closing the reference genome assembly by utilizing long-read PacBio sequencing in combination with a previously generated BioNano whole-genome optical map and Dovetail-HiRise map of chromatin interactions. 2) annotate the reference genome for the coding and noncoding transcript-isoforms and alternative splicing by full-length single-molecule sequencing. 3) annotate the genome for chromatin histone modifications and chromatin accessibility by integrating the RNA-seq, ATAC-seq, and ChIP-seq data across various tissues.

Publications

  • Type: Journal Articles Status: Accepted Year Published: 2022 Citation: Salem M, Rafet A-T, Ali A, Kenney B: Integrated Analyses of DNA Methylation and Gene Expression of Rainbow Trout Muscle under Variable Ploidy and Muscle Atrophy Conditions. Genes 2022, 13(7):1151.
  • Type: Journal Articles Status: Accepted Year Published: 2022 Citation: Ali A, Salem M: Genome-wide identification of antisense lncRNAs and their association with susceptibility to Flavobacterium psychrophilum in rainbow trout. Front Immunol 2022, 13:1050722.
  • Type: Journal Articles Status: Accepted Year Published: 2022 Citation: Ahmed RO, Ali A, Al-Tobasei R, Leeds T, Kenney B, Salem M: Weighted Single-Step GWAS Identifies Genes Influencing Fillet Color in Rainbow Trout. Genes (Basel) 2022, 13(8).


Progress 04/01/21 to 03/31/22

Outputs
Target Audience:Aquaculture, fish and animal sciencescientific community. Changes/Problems:We are making progress despite the pandemic and the supply chain disruption issues. What opportunities for training and professional development has the project provided?The project helped in training 2 postdocs, 2 graduate and 3 undergraduate students. How have the results been disseminated to communities of interest?The genome reference and transcriptome annotation have been published and are available through the NCBI databases. What do you plan to do during the next reporting period to accomplish the goals?Working on generating ATAC-Seq, Chip-Seq, and DNA methylationgenomeannotations.

Impacts
What was accomplished under these goals? A new genome reference was already published by Palt's group for the Arlee genetic line. New transcriptomeannotation and alternative splice variant database using the long-read sequencing technology have also been published. Current efforts are focused on generating ATAC-Seq and ChIP-Seq genome annotation.

Publications

  • Type: Journal Articles Status: Published Year Published: 2021 Citation: PacBio Iso-Seq improves the rainbow trout genome annotation and identifies alternative splicing associated with economically important phenotypes. Front. Genet. | doi: 10.3389/fgene.2021.683408


Progress 04/01/20 to 03/31/21

Outputs
Target Audience:Aquaculture and fish scientific community, 1 publication accepted. Training of 2 postdocs, 2 graduate and 1 undergraduate student. Changes/Problems:The pandemics caused major delays to this project but we have adapted. What opportunities for training and professional development has the project provided?The project helped intraining for 2 postdocs, 2 graduate and 1 undergraduate student. How have the results been disseminated to communities of interest?Salem's lab published a new transcriptome assembly and genome annotation usingPacBio long-read Iso-Seq technology. Palt's lab published the new genome reference from the Arlee line and it is now annotated and available in genome browsers at the NIH NCBI RefSeq and the Ensembl genome browser of the EBI (European Bioinformatics Institute). What do you plan to do during the next reporting period to accomplish the goals?Optimize ATAC-Seq,Chip-Seqprotocols to annotate the genome reference. Adding DNA methylation annotation to the genome reference. Use the new improvedgenome reference and it's annotations to identify structural and functional variants associated with important phenotypic variations.

Impacts
What was accomplished under these goals? A new assembly of the rainbow trout transcriptome (Swanson clonal line) has been published allowing identification ofalternative splicing associated with economically important phenotypes. Palti's lab released a new genome reference from a second genetic line (Arlee) allowing the identification of accurate structural variants and disease-related variants. PacBio DNA sequencing and generation of optical maps from the Swanson line and two additional distinct genetic lines (Whale Rock and Keithly Creek) are currently ongoing to generate 4 high-quality assemblies towards a pan-genome reference for rainbow trout.

Publications

  • Type: Journal Articles Status: Awaiting Publication Year Published: 2021 Citation: PacBio Iso-Seq improves the rainbow trout genome annotation and identifies alternative splicing associated with economically important phenotypes. Front. Genet. | doi: 10.3389/fgene.2021.683408
  • Type: Conference Papers and Presentations Status: Published Year Published: 2020 Citation: Palti, Y., Gao, G., Zhou, H., Salem, M. 2019. Improving the functional annotation of the rainbow trout genome. In Proceedings of the International Conference on Integrative Salmonid Biology. Fourth International Conference on Integrative Salmonid Biology, November 17, 2019, Edinburgh, Scotland,UK. P.No.Poster - 18.
  • Type: Journal Articles Status: Published Year Published: 2021 Citation: Gao, G., Magadan, S., Waldbieser, G.C., Youngblood, R.C., Wheeler, P.A., Scheffler, B.E., Thorgaard, G.H. & Palti, Y. (2021). A long reads-based de-novo assembly of the genome of the Arlee homozygous line reveals chromosomal rearrangements in rainbow trout. G3 Genes|Genomes|Genetics, 11. DOI 10.1093/g3journal/jkab052