Transcriptome-Analysis-of-Chromera(velia,-a-ChromeridAlga-Associated-with-Corals-of-the-Great-Barrier-Reef-Amin Mohamed1,2,4,6 , Vivian Cumbo2, Susanne Sprungala2,4, David Bourne3, Bette Willis2,5, David Miller2,4 1AIMS@JCU,*Australian*Ins3tute*of*Marine*Science,*School*of*Marine*and*Tropical*Biology,*James*Cook*University,*Townsville,*QLD* 4810,*Australia;*2ARC*Centre*of*Excellence*for*Coral*Reef*Studies,*James*Cook*University,*Townsville,*QLD*4810,*Australia* 3Australian*Ins3tute*of*Marine*Science*(AIMS),*PMB3,*Townsville,*QLD*4810,*Australia;* 4School*of*Pharmacy*and*Molecular*Sciences,* James*Cook*University,*Townsville,*QLD*4810,*Australia;* 5School*of*Marine*and*Tropical*Biology,*James*Cook*University,*Townsville,* QLD*4810,*Australia;*6Zoology*Department,*Faculty*of*Science,*Benha*University,*Benha*13518,*Egypt* BACKGROUND* • Chromera(velia(is*a*newly*discovered*algal*species*found* to*be*associated*with*corals* * • C.#velia#was*isolated*from*Magne<c-Island-at*the*GBR*in* 2012*from#Mon.pora(digitata#(Cumbo*et#al.,*2012)# * • C.#velia#is*closely*related*to*the*apicomplexanparasites,*but*also*related*to*the*photosynthe<cdinoflagellates*including*the*coral*symbiontSymbiodinium## • The*func<onal-role-of*C.#velia#in*the*coral*holobiont* is*currently*unknown-** Magne3c** Island* Fig.*1:*Loca3on*and*the*coral*species*Mon.pora#digitata#that** hosts*C.#velia** Fig.*2:*Nuclear*phylogeny*places*C.#velia#between*the* *Apicomplexa*and*the*dinoflagellates*Oborník*et*al.,*2012,*Pro3st* Fig.3:*Life*cycle*of*C.#velia:*(a)*predominant*coccoid*cells,*(b_c)* dividing*cells*(d_e)released*autospores*star3ng*a*new*cycle,*(f)* autospores*can*form*bi_flagellated*cells*in*favorable*condi3ons.* Oborník*et*al.,*2011,*Pro3st* PROJECT*AIM* * • To*generate*a*reference*transcriptome*assembly*of*C.#velia#isolated*from*the*coral*Mon.pora#digitata## ####at*the*GBR* * • To*Inves3gate*the*diversity*and*dynamics*of*C.#velia#transcripts*(mRNAs)* * → Genomic*insight*into*a*novel*chromerid*alga*via*iden3fying*genes*involved*in*various** *****physiological*and*metabolic*processes** → Differen3al*gene*expression*profiles*in*different*experimental*treatments* NEXT*STEPS* METHODOLOGY* I. Control-condi<on-II. Mixotrophic-condi<on-Experimental-- III. Mo<le/immo<le-Treatments-- IV. Heat-stress-- 3-replicates/- V. Heat-shocktreatment- De#novo#Transcriptome*Assembly*and*Analysis* Experimental-treatmentsSequencing* reads* (per*sample)** VII. Cold-shock-VIII. Darkness-- Abundance--es<ma<on- (all*samples)* Iden<fy-differen<ally-expressed-transcripts- Next*Genera3on*Sequencing*(NGS)* Total(RNA(isola.on(( ((((((((((((((((((((((((RNA(Quality(check(using#NanoDrop# #######################RNA(Integrity(check(using#Agilent#Bioanalyzer# (mRNA-enrichment* * -cDNA-library-construc<on*** Fig.*6.*Next_gen*DNA*sequencing*method:* * PCR*bridge*amplifica3on*and**Illumina* HiSeq*2500*sequencing*plaform* Illumina-highHthroughput-sequencing( * _ _ _ _ Combine*reads* Assembled* transcripts* VI. Cold-stress-- Fig.5:*C.*velia*cultures*growth*curve*and*experimental* treatments* Fig.*4:*Chromera#velia#cells* Reads*Normaliza3on** De(novo(assembly--Assembled* transcripts* KEGG-Metabolic-Pathways- Expression-paMerns-and-transcript-clusters- Adapted*from*Haas*et*al.,*2013*Nature*Protocols* References* Cumbo,*V.,*Baird,*A.#et#al.,*(2012):*Chromera#velia*is*Endosymbio3c*in*Larvae*of*the*Reef*Corals*Acropora#digi.fera#and*A.#tenuis.#Pro3st,*164:*237_244* Haas,*B.,*Papanicolaou,*A.,*et#al.,#(2013):*De#novo#transcript*sequence*reconstruc3on*from*RNA_seq*using*the*Trinity*plaform*for*reference*genera3on*and*analysis.*Nature*Protocols,*8:1494–1512* Oborník,*M.*Vancová,*M.,*Lai,*D.,*et#al.,#(2011):*Morphology*and*ultrastructure*of*mul3ple*life*cycle*stages*of*the*photosynthe3c*rela3ve*of*apicomplexa,*Chromera#velia.*Pro3st,*162(1):115_30* Oborník,*M.,*Modry,*D.,*Lukes,*M.,*et#al.,#(2012):*Morphology,*ultrastructure*and*life*cycle*of*Vitrella#brassicaformis#n.*sp.,*n.*gen.,*a*novel*chromerid*from*the*Great*Barrier*Reef.*Pro3st,*163(2):*306_323*
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