Usingopensource ROVtodevelopnovelwaysofcharacterisingthemarineenvironment

TeagueJ1,GrahamG2,AllenMJ31MRes AppliedMarineScience,PlymouthUniversity,PL48AA, 2SirAlisterHardyFoundation forOceanScience(SAHFOS),CitadelHill,PL12PB, 3 PlymouthMarineLabaratory (PML),Prospectplace,Plymouth,PL13DH

ARemotelyOperatedVehicle (ROV)isatetheredunderwaterrobotthatallowsthevehicle's operatortoremaininasafeenvironmentwhiletheequipment (ROV)worksinthepotentiallyhazardousenvironmentbelow.Theopensource ROV,(OpenROV),offersaversatilemechanism formaritimesurveying.

Theanalysis ofnucleicacidmaterialfrommarinesystemsiscurrentlycommonly achievedviaNiskin samplingandpreservationofnucleicacidsisdelayeduntilarrivalatthesurface.Thisprojectaimstodevelop aninsitunucleic acidsamplerwhichpreservesthepayloadandthereforeexpands thecapabilityoftheOpenROV platform.Focuswillbeonthecollectionofmarinebiomass fromseawaterandmetabolically‘freezing’theRNAprofileinsitu topreventdegradationandstress-induceddeviance uponbringingthesampletothesurface,withinasimple andcheapdeviceattachedtoanROV.Wewillaimtousethisdevicetomonitormicrobialcommunities in realtimeusingbasicmicroscopylighttoselectivelytargetspecificwaterbodies forsampling.

Thenucleic acidpreservationdevicewillbeconstructedusing variouscomponents suchasArduinoandRaspberrypiforprogrammablecontrolandaperistalticpumpforsampling coupledwithsensorsmonitoringdepth, temperatureandsalinity.

Introduction:

References: Blueplanet:seasoflife,television program,BritishBroadcastingCorporation(BBC),London, firstbroadcast:(12Sep2001).Census ofMarineLife.(2015).Niskin Bottles.[ONLINE]Availableat:http://www.coml.org/edu/tech/collect/niskin.htm.[ Accessed:23August2015].MarineTechnologySociety.(2012).What isanROV?.[ONLINE]Availableat:http://www.rov.org/rov_overview.cfm.[Accessed: 22August2015].ROV.org.(2015).WhatisanROV?.[ONLINE]Availableat:http://www.rov.org/rov_overview.cfm.[Accessed: 08January2015].Sochacki, F&kennedy,P.(2008).OCR ASBiologytextbook. Essex:Heinemann.pp.120.Teague,J.(2013).Producing andusinganROV(RemotelyOperatedVehicle)inbenthicvideosurveysaroundtheCornishcoast.FdSc MarineScience.FalmouthMarineSchool.

“Over60%ofourplanetiscoveredbywatermorethanamiledeep.Thedeepseaisthelargesthabitatonearthandislargelyunexplored.Morepeoplehavetraveledintospacethanhavetraveledtothedeepoceanrealm....”– (TheBluePlanetSeasofLife,2001).

Thisquoteoutlinestherequisiteforoceanexploration.Thisprojectaimstocontributetoouroceanicexplorationcapability.Itisenvisagedthatfuturemodificationofthedevicedevelopedinthisprojectwillbeutilised inthedeepseaenvironment.

TheProblem:mRNAenablescellstorespondtostressorenvironmentalchangesbyalteringexpressionofdifferentgenes,usuallywithinmilliseconds.(López-Maury, etal.2008).Therefore,currentsamplingstrategiesthatrelyonthecollectionandretrievalofwaterfromgreatdepthsarepotentiallyinaccurate:itcantakemanyminutestoretrievewater.Duringthistimethesampledcellswillberespondingtonewconditions theyhavebeenexposedtosuchaschanginglight,temperatureandpressure.Thisispartofthecellularstrategyforkeepinggeneexpression intunewithphysiologicalneedsdictatedbytheenvironment,promotingadaptationtoboth short- andlong-termenvironmentalchanges.(López-Maury, etal.2008).

TheSolution:Adevicethatcaninsitu:IntroduceacustomRNAprofilingsystemwiththecapabilitytopreservethesampleinsitu.EliminatingthearchaicpracticeofNiskin samplingandbringingittothe21st century.

Figure1,PML’sOpenROV fleet:Triton&Princess.Figure2,OpenROV “Triton”1stdeployment.Figure3,ProtoypeRNAProfilerBreadboard.Figure4,3DPrintedAttachmentrail&Niskin attachment.

Researchquestion:ArecurrentmethodsofnucleicacidpreservationinmarinesamplinggivingatruerepresentationofRNAprofilesfromdepth?

Aim:ToproduceawatersamplingpayloadfortheROVtoallowtheaccurateprofilingofRNAinthemarineenvironment.

Fieldtestingofthedevicewillhavetobeconductedmultipletimestoensurethatstatisticallyrobustanswerscanbeprovidedforthefollowingresearchquestions:v CanpreservedRNAbeextractedfromthedevice?v DoesthisRNAshowamoreaccuraterepresentationofasampleindirect

comparisontotraditionalNiskin methods?v CantheROVbeusedtoactivelytargetspecificareasofinterestinsteadofa

predefineddepthwithinthewaterbodythroughthemediumofmicrocopy?

ThenucleicacidsampleswillbearchivedatPMLat-80°Cforlongtermstorage.AnytranscriptomicdatawillbeloggedattheBritishOceanographicDataCentre(BODC)asperNERCnationalcapabilityguidelines.Thedesignofthedevicewillbe‘published’onOpenROVwebforumsandall3DprintfileswillbesharedviaThingiverse.com asperthespirtandethosoftheopensource aspectoftheproject.

(CurrentMechanism)