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RISCy Business: Development of a RNAi design and off-target prediction software

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OK I thought so and who we
are so I'm technicians I'm looking since 10 years in a plant research institute which is nearby about 200 kilometres from both and uh all focus is tool tools search for genes which are involved in plant-pathogen interactions interviews by leading into podium and you as a model system and you can see uh Barnett plants young body plans which are
infected this continuity which is a major problem in the field of science you have a lot of losses due to the and the try to find genes which are involved in this room interactions by some diverse genetics so how we can do so
you might know that the the genome of violence in the last so for instance for body it's almost twice because the size of the human gene and this means you have about 30 thousand genes so how to find genes which are involved in only in plant gene interaction so high you can do this and the use of force which is called an eye and interference this was discovered in the nineties and to give sort so they wanted to make opportunity arms
that show like you wouldn't want to make it dark and assault OK the the the DNA gene for the for the blue collar genes and selects at more of testing in the product and then we have between your palms feature back
and they did this and actually what happened was expected to the worst so they found that the
flowers have right what the here from these these types of of quantum and this gets ahead independent experiments but it's always turned out to be the same and it's still quite some years to find out that there's a mechanism behind this which is called so how does I works so if double stranded irony and distance the same which is very unusual so people saying that this mechanism is used tool defense against an I so if double standard and the investors said it's recognized by an
enzyme called instances here and on the dissect cuts the standard dominate into small pieces which are about 21 based which are called small interfering RNA will inspire me then and another enzyme detectors small I may take it more 1 of this event which is important for later so only 1 of the 2 strands states and data support this complex this this complex then is attached to the time a amount here and this the binding of this complex blocks the translation and you have no quality so if we go back to our overall quality of solution the natural quality in the in class which is their mother so I lost
my look so here you have the the
natural gene in the plant and the people and at more of the blue corner of without on this was like amazed by the plant 1st bind to the true nature of the gene and blocked
the translation for the core of the blue corner so you have no colonies so this sort looks so with this you can efficiently silenced and not bound you and took slavery since many years of people use the the Newman in bonds implanted in many years speeches and we also use it so we have developed a test is system where we can use this to it it's very efficient like what I think so that's I mean we Quality of austenitic so if you don't know what to is you switch
and you might even look what's happened OK some people use it ends but they have 2 forms of so the 1st problem is that you might not have missed target of other genes and the other problem is that not so every silencing is sufficient so how we can public that we wanted to look to to to make a software tool to predict and also to use this tool for the good design would and diffusion const so really we wanted to make
a software which can predict that's that's been the efficiency and also we wanted to test this office will mean made itself some years ago but in the last year we wanted to wanted data and to make real experiments and to see whether this condition is and of course you want to have a nice self-image from some windows and everybody can uses and combines all these features so what is the most
target so imagine that you have some you want to make a constant so you want to knock out the genes so you make artificially you can do this in the lab a double stranded dynamic contract and then you know you have to check whether you only target gene you want and not only another title as I said to have about 20 thousand genes in biology and so the chance that you might hit another target is quite high so as you can see here so again double stranded RNA split into small elements and then the user short alignment which is called ball title fight and about I tried to find all the matches from the small and filamentous against a large database for instance the biology of what human gene but there and if you will designed a good concept is should only find hints of the targets but now and then you might have off-target which is here which have of course much less so kids but they might be efficient enough also to to call science to have an off-target effects so you might not only hit your that you want but also other genes because for instance in body it's very common um that you have gene families which have the same sequence of a similar sequence and uh it might be that you also another gene from the same family impeachable modulation so it's important to to put the peace and to know what and maybe to to change of upon starts designed for so then the question
arises how much similarities acquired for being of target and use but that we a model which is called the molecular clock this states
that about everyone million and evolution 1 nucleotide change so if you have a common ancestor this has the sequence like this then after 1 years you have 1 major exchange so here for instance which is the see the to and here and to and into the and so on so after 2 million years we have another change and the use this tool design our
experiments so reconstructed serve 15 constants be made incident that deconstructs um whichever so we use 1 time between knowledge looks really looks at a stimulus on this center very good efficient constructs environments and tested and everything so we use our best to accommodate and make 15 construct syntactic constructs is increasing matching similarity so you have some sort disease only homogeneously have no change when you have 100 per cent identity and so the 1st 3 in smaller steps so 98 % 96 % on 290 % and then another and then you have this over the the short the decline of wild type and you expect that the hits you get a much of a decreasing as well as against the soul in the best case to 100 % of conformity heat tool on the scientist to the virus and decrease to to the audience so you have no you don't hit any your target
OK so we make these concepts in the test in our system and so unfortunate that have no time to explain what looks to his the because and you can see all the constructs and from of a sentence the concert no effect down to control so the control of common sense here and as much as strong the go dollars more the have been infected so would be the best and that must mean that the the the confidence significance to use dual about 5 4 experiments small we cannot afford cost expensive and available we and after 5 experiment do statistics and so on then is the what state significant or not so that means it's significant has no effect on the significant and you can see here it's very interesting that to performing useful to the united 2 % and in the state have an effect sold this silencing books and knocked out of you but starting from 90 % tool 50 million years or 2 . 2 % you have no effect on you know you lost it's not working anymore now the question raised by I same 59 hits of from Baltimore from the alignment so you have 59 assignments
which Mitchell Park so why you don't have an effect in and this the so the suspicion also what the people Nelson's longtime did not each assign a sufficient so you might have a assignments which are not efficient and as you can see here most likely and this and this 59 credit but there's simply not efficient enough so let that a tool to make the software we use a very
basic towards tool to to western music which artificial because people who did not know exactly what's going on and what needs to be sufficient for reuse stuff like agent with the beginning contingency content and temperature but in the last few months we wanted to the implementation the software and to have a better prediction of the society enables you to confirm it has always been so in the last couple of months many research paper the imperial beach desert a lot this efficiency especially in humans and it turned out that the term dynamic properties are very important also the structure of time and so things like many free energies of folding free ends or targets
accessibility so use this
and the 1st thing important thing is this concentration so you have this irony here and then they're going to we're not out and wants to send woman so how does this happen this happen like that and the sentence if for some the so the less they understand is easier the more free and open and this trend is taken from the from the this complex and once that is more so this goes of order of the minimum free energy it's Lazarus was less David will move from this side but now you might have components the 1 tend so it's might not complementarity what we emanating might have gotten the the wants and so this is our main is completely useless so this we included in our software to do more to make a strand selection into mothballs all of those a note that type of thing so we all
those calculations did after the 1st time over this selection to look only after this decision really heating to the attacked the 2nd thing that
just seems to be important and it's published money applicants there is a target site accessibility of 2 of the 5 so as you can see here that so this is the primary source of which is in in this sense it's not really like this you have a lot of folding so you can see here you have them you have complemented by means and looping sensor stuff and this is the secondary structure and this is the following
and this is the edge of the house looks in this sense is a very small and you see how much is folded into much is going on so it's not the flat line and you can see already here that some some places it's easier to target it then on so if you have fallen since the folding mentioned here it's very difficult to to assess for the assignment because all already done and so this part of the for instance might be much easier to access and you can calculate this in this
tool called and beautiful ever chosen was overbite his catalyst the local based their probability so it stays there this irony and the time of all all part of the time so we do this only on the part of the target of 2 m money because it's much easier to calculate than on this but just and it's kind the probability how much the mole accessibility these uh from the Sun to the to the target and you can do same for all for each base pair but also for an image so you can do this for the 1 man to man and intervals for the whole assignment 21 and people from that it seems to be the reason the design and awful interpretable as I've said if at least a base pairs of designer making can access soul and guess so if it's less so reflect tool toward tree-based then as you might be not enough to to stick to the rest of this and sold to the from so it's a good thing to take into to accommodate that is the entities 8 months of design image into the target and this
is something that will be talk our contacts so far for simplicity I so we checked for each of the 7 years who 1 and 2 and so on to force each consists of of simplicity i visual only the 8 evidence because that uniform descended
the most of the library if you take and you can follow constructive Richard so you can see here that the position of the sequence position so we have 500 base pairs of segments of the target and so here it's the shown accessibility is the 0 means decide for what his position of the target was most persistent and 1 means it's was very accessible and you can see here so if you'll remember that the old man years and could main has had an effect on the 5 million years so you can see here that the fall of the velum 18 years the 1 2 and 3 and 4 have quite a lot of
access to the site so this is the last constant is in effect and city has quite some existed the sites where for the 5 million constantly completely disappear the decision to our data so is this again efficient as you can see whole efficient and then is still an issue you make this quantity
if you look more closer to the 1st concept of the 1 you as the only years you actually can
see that you have some the just so for instance from the old to hundreds of base pairs who have not so many site from 100 to 200 250 sample from 200 to 300 you also have some and from the uninformative have not not only the fuel and format 5 from that you have quite a lot so this brings us to the idea to make 5 thousand construct which are called to which
we call the little constructs the only from the sound of a has match to the top so you have from want to harm match the 2 minutes on a target and act an incidence so it's not measured target then from 100 to 200 200 200 200 400 and 405 so we created these contracts and tested in our system
and that looks quite interesting I have to say that just stopped to look at the results before I went to the to the confidence just so you're not finished with this but this looks already quite intestines really got the strongest and most significant effect this 405 and a which you also corresponds label hazardous sites and have a lot of and they have no effect no significant effect is 300 to 400 then they also know excessive aside and for to and 100 it's the thing is not the only 1 that makes us in form amount the 1 2 0 crammed observing the constant but as a set the look on the data and also the shift to find here potential tend to define interested because this what was known from the human and is does not apply OK
then already want to come to enhance sold created
as software such as quot sci-fi this can be done downloaded and and at the moment on the news at the moment making the cost of forms like a tedious reached the we find can you can have a custom database you can find efficient siamese into Canada comment and the uh and find its target genes and this is just a
screenshot telescopes like so I
think it's very useful and I want wanted to make this talk just to show that also among and governments can do quite useful things and can also that's just it's except
from the communities publish an article about it and I hope you may like to the a the
whole process is going to be how much I have no idea how long stretches of sequence is of opinion and the database this if you have some piece of sound that you want to do something this is home on much data so it's a lot of the this is from the picture was us some yes it's nine-dimensional says and the boat I use uh database indexing to do this very fast so I'm not saying I don't know exactly the digits but it's good to be efficiently so 1 check from from the stuff it takes about hi seconds so it's quite fast hello and so I have 2 questions actually them so you have seen using by applying right do you use that to do your blast know what the article we the tried last beginning but it's also perform made that for sure what the sequence of so you have a lot of mistakes actually so it does not find all the assignments detection and there so if you look at on this graph from the 1st half you can see on the blue line so this was just the taken to align all the scientists is blossomed motives would have gaps in between cannot find all the scientists and that's what this vegetable by apartment just mentioned him because the use of force something and was like converting the the delivers complementing and opened fire stuff the fights and substance so this is most either using this model because you're saying in the beginning you're trying to find the all started to swell right the social part of this also yes it's all supported this signal about time OK so good getting piton uh the uh the credit sequence 21 household whatever things you like and then I take this as sequence and usable tight to to find the in my database which is also mating bold type in both of these reforms and the heat but you it's conceived here so this is actually it's would this is the target you want to keep but you might have targets of this also comes to do this and other gene and you might have few in this region you might have of target so this is done what this and this is like here just memory so this is so you know if you have if you have clearly and target which are the same and you group that you will find in each of the substrings on the on the product here so you have 80 but if you have a mismatch in between when it's you will find on the note that like and for this use all time and and just those wondering whether those targets I predictions they are based on the secondary structure in some yes you can so that just as a tool to look at the and we try to to find special many also legal for machine learning and stuff like this but insisted that no professional for combination in each time to develop thanks use the joint using bought was social readers and on this some short reads but I didn't get very hard to other room milligrams almost sort of worked it over the shortest novel what yes so you have you can and that you are going to suffer here you can pay your credit sequences we call it clearly the clients and then in Python I just did so I a tool for you you can choose the size of that what I meant are among the British here if you wanted to report use of whether grid against the topology or more central yes you can so this is a very big advantage of sci-fi so all was assist online and also for for dominant you cannot use a custom database so they usually for humans with a scientist usually very often used in human everything's adopted for humans but this is uses philosophy need to to to check against the normal or whatever so you can make a customized database and you can check against the In the same problem because you know the the sequencing of the human and the biology numbers ongoing and you might microphone makes the completely different sequence than last week so we need to be very flexible here is sort of the reporters used during armed published referring to me as a to sometimes you have a there you'll you have confidence because that's what we also don't go online at moment the working group was a short summary of the Florida AI researchers wondered how you can only if you like so you can download them and then and paste them and to use it but so we want to be very flexible this point because all the other stuff some interest so now the if so the
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Formale Metadaten

Titel RISCy Business: Development of a RNAi design and off-target prediction software
Serientitel EuroPython 2014
Teil 83
Anzahl der Teile 120
Autor Lück, Stefanie
Lizenz CC-Namensnennung 3.0 Unported:
Sie dürfen das Werk bzw. den Inhalt zu jedem legalen Zweck nutzen, verändern und in unveränderter oder veränderter Form vervielfältigen, verbreiten und öffentlich zugänglich machen, sofern Sie den Namen des Autors/Rechteinhabers in der von ihm festgelegten Weise nennen.
DOI 10.5446/20035
Herausgeber EuroPython
Erscheinungsjahr 2014
Sprache Englisch
Produktionsort Berlin

Inhaltliche Metadaten

Fachgebiet Informatik
Abstract Stefanie Lück - RISCy Business: Development of a RNAi design and off-target prediction software RNA interference (RNAi) is a biological mechanism for targeted inhibition of gene expression. It has also been used routinely to discover genes involved in the interaction of plants with pathogenic fungi. To minimize the miss-targeting of unrelated genes and to maximize the RNAi efficiency, we have developed a PyQt based cross- platform software tool called “si-Fi”. Our aim of the talk is to show that also hobby programmers can use Python in a very useful way.
Schlagwörter EuroPython Conference
EP 2014
EuroPython 2014

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