 Well, those of you who don't know what the Swiss Institute of Bioinformatics is, it's a big organization all across Switzerland with different groups doing surveys or research for bio, to federate bioinformatics in, in, across Switzerland. And so we are, the California group is a, is a joint group between the Swiss Institute of Bioinformatics and the University of Geneva. And we developed NexPROT since 2011. So where is NexPROT? What is this concept of a human database dedicated on human proteins? So we, we had this idea 10 years ago to create this specific database on top of Swiss PROT entries. So for those of you who are not very familiar with Uniprot, Swiss PROT, NexPROT, and all these landscape of protein databases, let me just explain you exactly the difference between all these PROT things. So you probably all heard about Uniprot. And Uniprot is a very, very big database on proteins from all organisms. The current release of Uniprot has about 200 millions of sequences from half a million species. From this very, very big corpus of sequences, some are manually reviewed. And now, so, so this manual reviewing is done on a restricted set of species and a restricted set of genes within those species. And in the current release of Uniprot, there are half a million of manually reviewed sequences, which which are available. And from these half a million manually reviewed entries, there are 20,000 entries for human genes. And in those entries in Swiss PROT, you find manually corrected and manually reviewed sequences and lots of biologically relevant information, such as the function of the protein, different features of the sequence, the presence of domains of post-transfessional modifications and so on. This information is extracted from the literature and also obtained by applying a number of sequence analysis tools on the sequences. But there are also a lot of other information on human proteins that is available on different resources. And the goal of NextPROT was to add this information that was not in the Swiss PROT entries and that was available in different resources at genomic level, transcriptomic level, proteomic level, protein-protein interactions and so on and so forth. So 10 years ago, we decided to take this very small subset of Swiss PROT, which is human protein entries, and add all what we could find in those entries and create really an integrated database on human proteins. So at present, we integrate, our main focus is, our main focus is our variant annotations and we integrate a lot of different polymorphism data from DBSNIP and Genomady. And lots of information about somatic mutations that appear in cancers and that come from the Cosmic Database. We also add information about the expression of genes in different tissues, and both at the mRNA level and at protein level. So it's antibody-based data, mainly. We add some information about protein-protein interactions, mainly from two sources, the intact database, and also a new set that was, that we integrated from any of the pharma, a company, a friend-based company. And we also have a strong focus on mass spectrometry data, and we integrate a lot of data from proteomic suppositories, such as massive or peptide atlas, to integrate information about peptide identification attached to the proteins. So all this information comes in NextProt. So just as a disclaimer, so because we build NextProt on SwissProt, we depend, our sequences are exactly the same that the one that you find in SwissProt. We don't have more sequences than SwissProt, we just add more information on the same sequences. And SwissProt and NextProt are considered to be complete in terms of human genome because they contain at least one sequence, one representative sequence, per known human gene. And in each entry, you find at least one of those sequences, and sometimes other splice isoforms, but it can happen that other isoforms exist and are in Uniprot, but not in the SwissProt of your part, but in Tremble. So these entries, you don't find them in SwissProt and you won't find them in NextProt. So if you are interested in having all the sequences described for human, then you have to turn to the very large corpus of Uniprot human sequences. So in the current release of NextProt, we have 20,353 entries, exactly the same number as SwissProt. And as you can see in this diagram, because we focus strictly on one species, which is human, we can have a coverage that is bigger than the one, larger than the one from SwissProt, in terms of protein-protein interactions, expression data, peptide data, and so on, that is not annotated in SwissProt. So we are more complete for human proteins than what you can find in SwissProt. And so, for example, we have nearly one billion different variants annotated in NextProt. And we have nearly 200,000 protein post-anxational modifications annotated. And we have two and a half million peptide identifications associated to human proteins that are annotated in NextProt. So it's a fairly big... NextProt offers a very big data set on human proteins. So NextProt has entries like SwissProt and the NextProt identifiers, in fact, are the same than the SwissProt ones, and we just add nx before the SwissProt identifier. And for each of the entry, we have different views, depending on what is your interest in the protein. And for each protein, for example, you have a function view, and then you have different views, medical, expression, interactions, localization, sequence, etc. And those views are displayed in the left menu, and you can change the views depending on the information that you are looking for. So here I show a screenshot of the function view of thyroredoxin, human thyroredoxin. And in this function view, you will find the summary of the function of the protein that is done by SwissProt curators. And after the summary, you have a list of go annotations that come from a variety of sources and that give for each annotation the associated metadata provenance and evidence of this annotation. In the sequence view, the sequence view allows you to tell, for example, for a particular protein, what variants are known, what protein post-translational modifications were described, and those variants and those PTMs, are they located in functional domains, are they affecting active sites, and so on. We build this sequence view in an interactive viewer, where you can find the sequence and all the annotations, the position annotations related to the sequence that are displayed in the upper part of the viewer. And then if you click on one of the features of interest, you can see it in the lower part of the viewer. And in this table, in the lower part of the viewer, you have all the metadata associated to a particular position annotation. And it's dynamically linked, and when you click either in the table or in the sequence display, then it highlights also the residue of a selected sequence on the sequence that is displayed at the right side of the screen. So the sequence view offers a dynamic representation of all position annotations, and gives a lot of annotated metadata associated to the data that is presented for the position annotations. And in addition, you have a search functionality in the feature viewer here, in which, for example, you can look for a particular genome AD variant, or a PTM, or any positional annotation, and that will direct you to the line in the feature viewer that corresponds to your query. And once you are in the line corresponding to your query, you have a link to the resource providing this annotation, and you have, of course, the details of the annotation. We also have an interaction, a view dedicated to protein-protein interactions. And in that view, you can find which parts of your protein of interest mediate interactions with other proteins. This is, for example, the interaction view of the HIP1 protein. And in this view, you can see with a similar display then for the sequence view, you see the sequence of the protein and you see the domains that support the interaction with other proteins. And like for the sequence view, you can navigate between those different viewers. We also have a view dedicated to the structure of the protein. And in this view, you can, for example, look if a particular variant will be externally accessible, or on the contrary, internal to the structure of the protein. So exactly like for the sequence view, you can select any positional annotation on the sequence viewer that will be highlighted on the feature table. And then below, you find a viewer of three-dimensional structure, and you can highlight any position of interest on any three-dimensional structure that is linked to the protein. We also have a view to look at the expression of the different genes, both at mRNA level and at protein level, with data coming from BG, which is another resource from SID, and from the human protein netlast, which is a resource giving data both at mRNA level by RNA-seq and a lot of antibody-based data. So we have put all this information in a single table that allows to compare the data at both mRNA level and protein levels for any tissue of interest. So as an important note for those of you who are developers, our sequence viewer, feature viewer, and the hierarchical heatmap table that is used for the expression view are all JavaScript components and can be freely reused if you are interested, and we show source code and examples in our GitHub repository. If you are a programmer and you want to retrieve next-pot contents, we can also use our API. And so in the next-pot website, we have this API with a help to how to use it, but basically you can retrieve any part of the entry of interest quite in a quite easy way. And I encourage you to have a look and try and to join to the specific hands-on breakout session that will take place later this afternoon on the API. Well, because next-pot has a lot of information specific to human proteins, it has been selected as the reference knowledge base for the human protein project of the Human Proteomorganization, QPo. So for those of you who don't know about this project, it's a project that was launched exactly 10 years ago on the 23rd of September, 10 years ago, and whose mission is to use mass spectrometry to validate all the predicted human genes. And so it federates about more than 1000 researchers worldwide, and each team tries to have really a credible identification for all the human genes. And because next-pot is the reference knowledge base for this project, it has to integrate all the data coming from this project, provide dedicated viewers and tools for this data, provide some export format that can be used by mass spectrometry researchers, and monitor the progress of the project and provide annual metrics. And one of the hands-on sessions will be specifically dedicated to proteomics data and tools in next-pot. So as I mentioned before, one of the important goals of this project is to validate the existence of all the human genes by mass spectrometry. And to validate those proteins, the first thing was to establish some criteria of validation of those proteins. And iteration after iteration, the HPP decided that one entry, one protein, was upgraded as validated by mass spectrometry, if at least two unique peptides that are non-nested are reported by one major repository. So what our role in next-pot is to integrate all the data that comes from the community at large and by HPP investigators in particular, who also submit their data individually to different mass spectrometry repositories. Then this data is re-analysed by the major global mass spectrometry databases that are peptided at NASA and massive, and that guarantee that the peptide identifications are of good quality. And then we as next-pot as a final resource for this project, we integrate all the peptides that have been validated by peptides. And we count if at least two of those peptides map to a particular protein and come from a single resource, either peptide Atlas or massive. And if yes, then the protein is validated at protein level and is validated. Next-pot also integrates data from another major partner of the HPP project, which is the human protein Atlas. But at the moment, antibody-based data that is integrated in the next-pot is not used to validate the existence of protein because of potential problems of antibody specificity. But we are thinking on how to also use antibody-based data to validate the existence of human proteins. So once we have integrated all this data, we compute a score of validation of the protein and all the proteins that have been confidently identified have a score of what we call E1. And we have other scores for protein not having sufficient credible evidence for their existence. And we have a complicated system of P2, P3, P4 categories for proteins that have good evidence that they should exist but have not been validated at protein level. And we also have a specific category which is called PE5 for entries for which we doubt that they really are protein coding and that probably correspond to pseudogenes or other elements of the genomes that are not real, that we don't think are real proteins. And based on all these counts and all these scores attributed to each protein, every year we estimate the number of proteins which still have to be tracked and validated by mass patronage. And in the last releases of next plot, there are still about 10% of the human protein which completely lacks any identification at protein level. So all this proteomics data that comes either from HPP researchers or from any other mass spectrometry labs are integrated and are displayed in a dedicated view that it's called the proteomics view. So in this view, it's built with the same principle that the sequence view. And in that view, you can find which peptides for a given protein have been found and in which tissue or which cell line. You can find if there is an antibody available for this protein and data available at the human protein atlas. And we also integrate data from SRM atlas, which is an atlas of synthetic peptides. And because of that you can immediately see what peptides you can use if you want to do some targeted proteomics studies. So all those identified peptides are represented here on the sequence view. And as for the sequence view, you can select any of them and have the details of the identification of the provenance of the quality of your peptide of interest. One important thing about those peptides are that we have to check that a mapping of one peptide to a protein is unique to this protein or can be found in other proteins. Because that's a strong criteria to be sure that those peptides can be used to validate the existence of the protein. So for each of the peptides that are reported for an entry, we report if they are uniquely mapping to this protein or not uniquely mapping to this protein. And of course we describe where the peptides have been identified. And if we have link to repositories where they have been found, we provide those links and then you can access directly to the spectra or to more technical details on the peptide. So as I just mentioned, one important aspect of peptide data integration in Nexprat is to validate the uniqueness of those matches from peptides to proteins. And to do that we developed a specific tool that is called the Nexprat peptide uniqueness checker that we use internally to check the uniqueness of all the peptides that we integrate. But also that we also proposed for the community to use to analyze their own list of identified peptides within a protein mix experiment. So the tool is accessible on the Nexprat website and basically it works as follows. So you first enter your list of peptides or you upload a file with all your peptides. You launch the query and then you will, for each of your peptides, you will see if it is uniquely mapping to an entry or if there are additional mappings. Both if you want to consider all the variants, the genomic variants known and described in Nexprat, or without taking polymorphisms into account, you have two choices. And so for example, here the second peptide is uniquely mapping to this entry when you don't take into account variants. But if you take into account all the variants that are known and described in Nexprat, this peptide loses its uniqueness. And of course you have other peptides that map to a number of entries. And even if you don't take into account any polymorphism, it will map to plenty of entries. And of course these results can be downloaded. You can have them in an Excel format and which is more convenient than looking at the results on this viewer. We also developed another tool that may be of interest for the protein missions, is the Nexprat protein digestion tool. So that works when you really want to focus on a particular protein and you want to design a specific experiment to identify this protein. And you want to select the proper enzyme to use for your mass spectrometry study. So in that case, you just query for your protein of interest. You give the length of the peptide that you want to get at the end of the cleavage. And then the tool will perform a series of theoretical digestions with different enzymes and will provide you with the counts of peptides and the uniqueness of those peptides. So you can predict if a given protein will work with a classical trypsin-based experiment or if you have to turn to another enzyme to be sure that at the end you will get unique peptides and you can validate your identifications. So this tool is also available in the Nexprat platform. So now we have seen how to look at data for one given entry. But another thing that is really useful to do with Nexprat is to search for information for global information and not only to start with one single entry. And there are different ways to search for information in Nexprat. The very simple one is the simple search in Nexprat which is a Google-like search in which you can search for proteins, of course, but you can also search for publications for all the publications that were used to annotate data that is found in Nexprat. And you can also search for terms that have been used in Nexprat. So that's quite large as a query, but it's not super precise. You cannot do really a combination of different queries. You cannot look for particular proteins in a particular context and so on. It's really like Google. So here I put an example. If you look for liver in the terms that are loaded in Nexprat, you see that you have a number of terms that cover, of course, the anatomy, but also some phenotypes or some go terms, etc. So that's quite convenient if you want to do a quick search. But that's not, in most cases, it's not really as precise as desired. So this is why we designed advanced tools really to make precise queries in Nexprat. And those tools are based on a technology that is in a language that is called Sparkle. Sparkle is a semantic query language that is able to retrieve and manipulate data that is stored in a format that is called RDF. So without going into a lot of details, a RDF DACA model is based on statements that are known as triples and that are, that have a subject, a predicate and an object. For example, the sky has the color blue. And we will detail that a bit more in the practicals after this lecture. So the Nexprat has a RDF-based model. Of course, it's a bit more complicated than the sky has the color blue. And our RDF data model looks like that. So in fact, for each of the Nexprat entries, you have different splice isoforms and all the annotations of expressions, sub-cellular locations, interactions, all the positional annotations are linked, attached to each of the isoforms present in the entry. So our model is isoform semantic. And for each isoform, you have a wide variety of annotations. And for each of these annotations, we don't use free text because it's not really standardized and interoperable. But we use controlled vocabularies as much as possible. And different terms and different ontologies that also are available on our ftp site. And we try to document for each of our annotation to document always the quality of the annotation, the provenance, and the detection method and all the metadata that we can capture to allow a full traceability of all our annotations. And so our Sparkle-based tools make use of this very rich data model and allow we to go far into exploration on Nexprat data. So Sparkle queries always consist of variables at the beginning. Then you have the query per se. And in some cases, you can modify the query using some modifiers. And so they look like that. This is one very simple Sparkle query where I am asking for all the proteins phosphorylated and located in the site of Plasm. This query cannot be done with a simple search. So you have to turn to the admin search. And the query is quite simple. You want entries where at least one isoform because all the annotations are linked to the isoform are phosphorylated. So we have this keyword here, which is the keyword associated with all phosphorylations. And we'll be located in the site of Plasm and or any subpart of the site of Plasm in Sparkle that is written like that. So I completely understand that when you don't know about Sparkle, it's quite difficult to write this kind of query. So we have made different queries that you can find on the Nexprat websites and that help the user to write their own queries. So in the advanced search of Nexprat, you have this interface which allows to retrieve lists of protein entries with Sparkle queries. Here is an example on how to look for those proteins missing evidence of detection of protein never. And you see that you can immediately get the complete list of them in one query. And as I mentioned before, to help, there is a pre-made list of queries available. But we also developed an even more advanced mode of query of Nexprat, which is called Snorker. And in that particular tool, you can not only retrieve lists of proteins, but you can retrieve absolutely any data that is in Nexprat. Not only proteins, but variants or families or any item that you are interested in and that is in Nexprat. And for example, you can retrieve all the variants that have a frequency of more than 0.1 and that will affect PTM sites. So the queries are generally a little more complex and you have the results in different formats. But for example, as a table here, and here with this example of the frequent variants that affect known PTM sites, you have immediately your list of results showing that, for example, here, this protein has a blank oscillation here and at the same position a variant from N to S with a very high frequency. So that's super convenient to retrieve complex sets of data from Nexprat in a format that is completely up to you, because in the variables of your query, you can ask for absolutely any item that is in Nexprat. And also for the Snorker, we have made some queries and we display them and we encourage people to use them and to customize them depending on their interest. And we just published a tutorial on how to use these different Sparkle tools to retrieve data in it. And of course, we also have detailed help on the different RDF entities that are in Nexprat and that help to write Sparkle queries. Nexprat contains a lot of things, but does not contain everything on human proteins. I mentioned that it contains mass spectrometry data, antibody-based data, RNA sequencing data, PTMs, variants, protein-protein interactions, functional annotations, which is already a lot, but it does not have any information about phylogenetic information, about data on model organisms, about pharmacology data, about clinical proteogen data, about structural data itself, or about interactions with pathogens and lots of other things. But the beauty of Sparkle-based tools is that in one query, you can not only query the content of the resource that you are querying, at the moment you are querying, but you can write queries that federate the corpus of data of one resource with other resources that are semantically compatible. And that is called federated Sparkle queries. And we already worked with a number of resources that have a Sparkle endpoint and that are semantically compatible with Nexprat. And in one query, you can really query both in the protein world and, for example, in the small molecule world or in the pathway resources and so on. And we have written lots of examples of such very powerful federated queries. And from time to time, we publish them in our website as examples. And we encourage people to give us ideas on queries and on resources to federate with Nexprat. And among the last queries that we released, we published some queries, for example, related to coronaviruses. And you can find here proteins involved in coronaviruses pathways that have medical information from other repositories or that have drug information from other repositories and so on. Last but not least, Nexprat integrates its own data, federates its data with other resources. Nexprat also provides their tools for the community, such as the proteomics tools. But Nexprat also wants to host tools that are developed by the community. And in the current release, we have two tools that are completely developed by third parties. And we just create links between Nexprat and those tools. And we allow our users to seamlessly navigate between Nexprat and these other tools. And those community tools can be found in the menu at the bottom of each entry. And if you are a developer and you have a tool, and you would like to distribute it through Nexprat, then Just Contact Us is quite easy to plug any tool to Nexprat. So don't hesitate to come and talk to us. So with that, I think I've gone to a pretty wide tour of Nexprat functionalities. And I wanted to thank my colleagues from the Geneva group, Amos Bayrod, who is co-directing the group with me. Paola Dweck, who is a bio curator in the team, guaranteeing that all the control vocabularies are properly used and that all the data is so she's deciding all the thresholds of quality for all the data that is integrated in Nexprat. We have a wonderful team of developers, Pierre-André Michel Vincenzo-Daponte, Alain Gâteau, Valentin Régia Laval, and Kassoul Samaras-Md. And Monique Sanne, who will be with me for the practicals and who guarantees the overall quality of the resource and who is really keen on answering feedback from the users. And with that, I thank you for your attention and I will be happy to answer any questions. Thank you very much, Lydie. Thanks a lot. And a big applause for you. Before we go on with the questions, please, you can use the chat to put your questions. You can also take the microphone and ask directly also to Lydie or use the Google doc that you have received also. But before we go into that, I would like to launch a poll to ask you three questions if you can. And so the questions that you see on your screen, while that will be wonderful is three quick questions. And in the meanwhile, while you are going through that, let's open then the chat or the microphones for your questions to Lydie. Any questions? Don't be shy. Okay, there's one short question. Yes, on the chat. Please go ahead or you can also put your microphone on. Hey, as you wish. Hi. Hello. Can you hear me? Yes. Oh, yeah. I have a short question on the criteria of missing 14. You said a single source. You said a single source of samples. What does it mean, the single source? I think a man is a single source or data sets means a single source. In fact, all the data that is submitted to individually to the different mass spectrometry repositories are reanalyzed either by peptide atlas and by messy. And the HPP decided to validate a protein. If you can find two peptides for a single sequence that come either from peptide atlas or from massive, but you cannot have one peptide from peptide atlas and one peptide from massive, that does not work because the first discovery rate would be too high for that. So I don't know if it's clear, but it's not that the two peptides have to be found in the exact same individual study, but no, but it has to be found in the same global reanalysis of all the mass spectrometry data that is submitted at a given time. So you mean the single source means in single data set, right? In a single big data set coming from the reanalysis of all the data sets. All the data sets. Okay. So what types? Individual data set. No, all the individual data sets are put in a global pot and then that last reanalysis all those data sets with a pipeline and massive does the same with a bit different individual data sets and each received from peptide atlas one very big data set and we receive from massive another very big data set. To qualify a protein has to have two peptides either from peptide atlas or from massive, but not one from each. Okay. Thank you. Thank you very much. There's another question also from Anna Garcia-Martin. Thank you for the lecture. Really great. I wonder if in a short period of time next brought we also compile information regarding bacterial proteins. Oh, that's not in our plans, but that would be very interesting. So no, next product is really built to focus on human proteins and we don't have at least in the in the midterm future. We don't have any plan to capture information on bacterial proteins, but what would we would like to do would be to have information on which human proteins can bind bacterial proteins and to have information about protein protein interactions between human and any kind of bacteria, but we don't want to be a database focusing. We don't have the resource to accommodate and to annotate bacterial proteins at the moment. Thank you very much. There is another question from Francesco Varato. How many researchers do you think or know use the next plot platform currently and how many from the EPFL? Good question. How many researchers so in the in the past years we have focused a lot on the proteomics community. And so I know that nearly all the HPP investigators use next plot because they also because they are required to use it to report their results. And that means more than 1000 people have to use next plot and I hope are happy with what we propose. Then we have also some users interested in rare disease variant in Europe because we are part of a network of teams working on rare disease genome annotations. But it's less than the potential users. And then specifically how many users from the EPFL I have no clue, but I could check the logs. Or I would be happy to hear if any EPFL lab is using us and would like to have and would have specific questions or specific fields of interest that we could help. If we have anyone on the call from the EPFL they can put that on the chat already. Yeah. So the next question from Echeverry. Amazing overview and amazing integration of massive information. I have a question regarding PTM mapping. PTM mapping. I just went through some proteins and saw mainly only mapping of fossil peptides. What is the evidence necessary to map other types of less common modifications like carbonylation or any other PTMs? Yeah. So we have different sources of PTM information in next plot. So for phosphorylation we rely on the phospho peptides that are analyzed by peptide atlas. So we receive them in one very big data set every two years depending on peptide atlas. We have further some modifications that need to be to be studied really at the level of one protein and that are not amenable to mass spectrometry study. We rely on the Swiss plot annotations and we integrate all the PTMs annotated by Swiss plot. And in between for all the other. So for glycosylation we integrate information from GlyConnect which is another resource from SIB and which annotates glycosylation based on different techniques, mass spectrometry but not only mass spectrometry. And for other modifications like ubiquitination or acetylation and so on we have our own annotation from mass spectrometry data. And so we take papers and we have our criteria to integrate this information. And so you can find ubiquitination sites, methylation sites and acetylation sites from a number of studies. But now we so we used to do that a lot in the past and now that the team is a bit reduced we don't have sufficient resource to cope with all the new studies coming with new large-scale PTM annotations. So we are currently trying to find solutions to overcome that and to be really up to date with all the PTM information which are non-phosphate. We are thinking we are we want to collaborate also maybe with a peptide atlas for other PTM annotations. We will see. But at the moment we have three or four different sources of PTMs not only phosphate. Thank you very much from David Lyon. Do you collect data on peptide protein abundances and how do you integrate data from various sources and experiments? Not yet. We don't we don't have any data on quantitative proteins yet. It is something that that is also under reflection and I would be happy to to have inputs on people involved in that field to to know how to integrate different abundance different results of different abundances for the same tissue etc. It's not trivial to integrate these kind of results but it's definitely very interesting to do. It's in our plan but we don't know yet when we will be able to have that. Thank you. From Geet Vora in terms of data what annotations and data are completely unique to next plot compared to Uniprot? Compared to Uniprot, in Uniprot you don't have mass spectrometry data integrated so all the peptides are not found in Uniprot. The expression data the details of expression data is not found in Uniprot. The phosphorylation sites from peptide atlas and the one in the PTM sites that we re-annotated based on proteomics papers are not in Uniprot. The last big dataset of protein-protein interactions from a new pharma is specific to next plot and not in Uniprot. Maybe there are other things that do not come to my mind just now but there are a number of things that are not in Uniprot. Thank you very much. Another question from Chia Wei, for peptide uniqueness checker what does it check exactly? Does it check the sequence peptide uniqueness to the whole human proteomic database or only isomers and how often does next plot update every three months or so? The uniqueness checker checks a peptide sequence against all the sequences that are in next plot so that means all the isoform sequences that are described in next plot which are the same than the ones that are in Swiss plot. That does not check the uniqueness of sequences against all possible human proteins that are in tremble and that are not been manually reviewed and integrated in Swiss plot and next plot. So it checks within the next plot view of the human proteome but it takes into account all the variations that have been annotated so all the polymorphisms all the somatic mutations and so on so are taken into account so that means that if a peptide if you have a variant that will change the sequence and that the peptide would match the sequence if it has this variation then the uniqueness checker is able to to to detect that this event. So it can never be complete because there are always new sequences coming and so on and we cannot guarantee that we have all the the human sequences existing and reported but we do our best to to have a representative human carefully manually reviewed that asset and for the second question about the updates of next plot so we try to to have at least two major updates per year and in some years we managed to have three so it's a bit less frequently updated than Swiss plot the new plot that makes an update every two months. Thank you very much another question from Espoir Cabanga thank you for the clear presentation this next plot uses any deep machine learning for artificial intelligence algorithms in either of its options or tools like for instance in the peptide uniqueness checker. No we don't it's the peptide uniqueness checker is not does not use this kind of algorithms and but we are not against developing such tools but we don't have any use case yet where we thought it would be relevant but I'm open to any suggestion and any collaboration in that field because there are plenty of things to do. Thank you from Michela Dalangelo thank you for the interesting talk are you planning to add phylogenetic information? We would love to it's not that easy because the different phylogenetic databases often have different results depending on the methods that they use and depending on the the extent of species that they are covering and so this information is a bit difficult to reconcile and we are currently testing different different methods and yeah but that's one of the things that we would like to add in quite near future