On the path to reference genomes for all biodiversity: lessons learned and laboratory protocols created in the Sanger Tree of Life core laboratory over the first 2000 species
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Abstract
Since its inception in 2019, the Tree of Life programme at the Wellcome Sanger Institute has released high-quality, chromosomally-resolved reference genome assemblies for over 2000 species. Tree of Life has at its core multiple teams, each of which are responsible for key components of the ‘genome engine’. One of these teams is the Tree of Life core laboratory, which is responsible for processing tissues across a wide range of species into high quality, high molecular weight DNA and intact RNA, and preparing tissues for Hi-C. Here, we detail the different workflows we have developed to successfully process a wide variety of species, covering plants, fungi, chordates, protists, arthropods, meiofauna and other metazoa. We summarise our success rates and describe how to best apply and combine the suite of current protocols, which are all publicly available at protocols.io.
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AbstractSince its inception in 2019, the Tree of Life programme at the Wellcome Sanger Institute has released high-quality, chromosomally-resolved reference genome assemblies for over 2000 species. Tree of Life has at its core multiple teams, each of which are responsible for key components of the ‘genome engine’. One of these teams is the Tree of Life core laboratory, which is responsible for processing tissues across a wide range of species into high quality, high molecular weight DNA and intact RNA, and preparing tissues for Hi-C. Here, we detail the different workflows we have developed to successfully process a wide variety of species, covering plants, fungi, chordates, protists, arthropods, meiofauna and other metazoa. We summarise our success rates and describe how to best apply and combine the suite of current protocols, which …
AbstractSince its inception in 2019, the Tree of Life programme at the Wellcome Sanger Institute has released high-quality, chromosomally-resolved reference genome assemblies for over 2000 species. Tree of Life has at its core multiple teams, each of which are responsible for key components of the ‘genome engine’. One of these teams is the Tree of Life core laboratory, which is responsible for processing tissues across a wide range of species into high quality, high molecular weight DNA and intact RNA, and preparing tissues for Hi-C. Here, we detail the different workflows we have developed to successfully process a wide variety of species, covering plants, fungi, chordates, protists, arthropods, meiofauna and other metazoa. We summarise our success rates and describe how to best apply and combine the suite of current protocols, which are all publicly available at protocols.io.
This work has been peer reviewed in GigaScience (see https://doi.org/10.1093/gigascience/giaf119), which carries out open, named peer-review. These reviews are published under a CC-BY 4.0 license and were as follows:
Reviewer 2: Lars Podsiadlowski
The Authors provide a profound overview over their aim to generate genome information for a wide range of species in the tree of life project. As a scientist with hands on experience on genome sequencing, I greatly appreciated all the information here, especially detail on the differences experienced with different taxa, as this is probably the most important lesson here, that there is high variation and strategies must be adapted to that. I am also happy that many of the approaches are also available as detailed online protocols, which really helps a lot in practical work. The selected examples of size profiles also give a good impression on what differences can be expected, e.g. with different extraction methods applied to the same species. Although detailed, I think that the authors provide a lot of relevant information here and would not change that. I did also not spot any errors or flaws in the text.
One thing that might be changed is the title. From first reading it I expected to hear also about assembly strategies, as well as some comparisons and oddities of the yielded genomes. It is great to have the manuscript as it is, but I like to see it better reflected in the title that the main focus here is on the wet lab part, especially the extraction of good quality DNA/RNA.
I have some issues with the figures: Fig. 7: there is no mention in the legend about the y-axis scale - I assume from the text that it refers to Gigabases? Figs. 8,9, 11-15: It is a bit confusing until I realised the log scale of the numbers. I would prefer to see it not with a log scale, but in a similar way as Fig. 6, with percentages on display, and an accompanying species number somewhere on the side. In the way it is shown now, the failed proportion looks so small and gives a wrong impression. Maybe overthink the colors, I would prefer another color for the Pass ULI, which is more similar in tone with Pass, because at the moment pass ULI and fail are similar in tone and brightness and appear as being opposed to the green "pass", while the difference between "fail" and the rest should be more pronounced in my view.
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AbstractSince its inception in 2019, the Tree of Life programme at the Wellcome Sanger Institute has released high-quality, chromosomally-resolved reference genome assemblies for over 2000 species. Tree of Life has at its core multiple teams, each of which are responsible for key components of the ‘genome engine’. One of these teams is the Tree of Life core laboratory, which is responsible for processing tissues across a wide range of species into high quality, high molecular weight DNA and intact RNA, and preparing tissues for Hi-C. Here, we detail the different workflows we have developed to successfully process a wide variety of species, covering plants, fungi, chordates, protists, arthropods, meiofauna and other metazoa. We summarise our success rates and describe how to best apply and combine the suite of current protocols, which …
AbstractSince its inception in 2019, the Tree of Life programme at the Wellcome Sanger Institute has released high-quality, chromosomally-resolved reference genome assemblies for over 2000 species. Tree of Life has at its core multiple teams, each of which are responsible for key components of the ‘genome engine’. One of these teams is the Tree of Life core laboratory, which is responsible for processing tissues across a wide range of species into high quality, high molecular weight DNA and intact RNA, and preparing tissues for Hi-C. Here, we detail the different workflows we have developed to successfully process a wide variety of species, covering plants, fungi, chordates, protists, arthropods, meiofauna and other metazoa. We summarise our success rates and describe how to best apply and combine the suite of current protocols, which are all publicly available at protocols.io.
This work has been peer reviewed in GigaScience (see https://doi.org/10.1093/gigascience/giaf119), which carries out open, named peer-review. These reviews are published under a CC-BY 4.0 license and were as follows:
Reviewer 1: Yuan Deng
The manuscript focuses on the entire experimental processes involved in the generation of high-quality genomes and proposes a set of standardized and modular experimental process protocols. The innovation of these protocols is that they can be flexibly combined according to different taxa, tissue types and sample quality, which greatly improves the flexibility and efficiency of the experiment and provides a reference experimental process for researchers in this field to follow. The manuscript also explore the specific challenges and solutions of different taxa in the experimental procedure of sample processing, DNA extraction, shearing, cleaning, Hi-C and RNA extraction, providing valuable guidance for future research. Meanwhile, the manuscript reviews the experimental protocols for the production of genome data of more than 2,000 species, which is in line with the journal's focus on biological big data. Therefore, I consider the subject matter and content of this work are appropriate for publishing in this journal. I only have some minor requests for revision:
1.Sample processing: (1) Sampling of rare and endangered species: for such a large-scale study of the "Tree of Life", it is bound to involve some species that are difficult to obtain conventional tissues, therefore the manuscript may include a section on how to select suitable tissues for subsequent experiments, especially for rare species. And is it possible to provide a prioritized list of tissues selection based on the difficulty of extracting high-quality DNA? (2) Processing and extraction of unconventional tissues: accordingly, it is recommended to add content regarding sample processing and extraction procedures for unconventional tissues, e.g., any particular methods to improve the quality of DNA extraction. (3) Sample contamination problem is often overlooked yet critical: how to reduce sample contamination problems in large-scale sample processing and other experimental processes? How to exclude sample or experimental contamination from data?
2.Analyzing method limitations: while the manuscript mentions some challenges that may be encountered in the processing of samples from various taxa, there is little discussion on the limitations of those experimental methods. It is recommended to expand the content of the limitations of the methods, such as some methods may not work well for certain types of samples, or some steps may have factors that affect the accuracy of the results, so that readers can have a more comprehensive understanding of the scope of application and potential problems of the method.
3.The manuscript is currently organized according to the experimental procedures, but some of the more relevant components could probably be consolidated to reduce redundant information and improve the readability. The authors studied the experimental conditions for different taxa in long read sequencing and Hi-C library preparation, but fail to emphasize their relevance in the introduction.
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