Chapter 10 Two Factor Designs - Single-sized Experimental units - CR and RCB designs Contents 10.1 Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 527 10.1.1 Treatmentstructure . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 528 10.1.2 Experimentalunitstructure . . . . . . . . . . . . . . . . . . . . . . . . . . . 536 10.1.3 Randomizationstructure . . . . . . . . . . . . . . . . . . . . . . . . . . . . 538 10.1.4 Puttingthethreestructurestogether . . . . . . . . . . . . . . . . . . . . . . . 539 10.1.5 Balance . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 539 10.1.6 Fixedorrandomeffects . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 540 10.1.7 Assumptions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 541 10.1.8 Generalcomments . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 542 10.2 Example-Effectofphoto-periodandtemperatureongonadosomaticindex-CRD 543 10.2.1 Designissues . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 543 10.2.2 Preliminarysummarystatistics . . . . . . . . . . . . . . . . . . . . . . . . . 544 10.2.3 Thestatisticalmodel . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 548 10.2.4 Fittingthemodel . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 549 10.2.5 Hypothesistestingandestimation . . . . . . . . . . . . . . . . . . . . . . . . 549 10.2.6 Modelassessment . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 554 10.2.7 Unbalanceddata. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 555 10.3 Example-Effectofsexandspeciesuponchemicaluptake-CRD. . . . . . . . . . 555 10.3.1 Designissues . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 557 10.3.2 Preliminarysummarystatistics . . . . . . . . . . . . . . . . . . . . . . . . . 557 10.3.3 Thestatisticalmodel . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 559 10.3.4 Fittingthemodel . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 559 10.4 Powerandsamplesizefortwo-factorCRD . . . . . . . . . . . . . . . . . . . . . . 564 10.5 Unbalanceddata-Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . 567 10.6 Example-Streamresidencetime-UnbalanceddatainaCRD . . . . . . . . . . . 569 10.6.1 Preliminarysummarystatistics . . . . . . . . . . . . . . . . . . . . . . . . . 571 10.6.2 TheStatisticalModel . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 572 10.6.3 Hypothesistestingandestimation . . . . . . . . . . . . . . . . . . . . . . . . 573 526 CHAPTER10. TWOFACTORDESIGNS-SINGLE-SIZEDEXPERIMENTALUNITS-CRAND RCBDESIGNS 10.6.4 Powerandsamplesize . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 579 10.7 Example-Energyconsumptioninpocketmice-UnbalanceddatainaCRD . . . 579 10.7.1 Designissues . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 580 10.7.2 Preliminarysummarystatistics . . . . . . . . . . . . . . . . . . . . . . . . . 580 10.7.3 Thestatisticalmodel . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 582 10.7.4 Fittingthemodel . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 582 10.7.5 Hypothesistestingandestimation . . . . . . . . . . . . . . . . . . . . . . . . 583 10.7.6 Adjustingforunequalvariances? . . . . . . . . . . . . . . . . . . . . . . . . 587 10.8 Example:Use-DependentInactivationinSodiumChannelBetaSubunitMutation -BPK . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 587 10.8.1 Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 587 10.8.2 Experimentalprotocol. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 588 10.8.3 Analysis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 588 10.9 Blockingintwo-factorCRDdesigns . . . . . . . . . . . . . . . . . . . . . . . . . 598 10.10FAQ . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 598 10.10.1Howtodeterminesamplesizeintwo-factordesigns . . . . . . . . . . . . . . 598 10.10.2Whatisthedifferencebetweena‘block’anda‘factor’? . . . . . . . . . . . . 599 10.10.3Ifthereisevidenceofaninteraction,doestheanalysisstopthere? . . . . . . . 599 10.10.4WhenshouldyouuserawmeansorLSmeans? . . . . . . . . . . . . . . . . . 601 Thesuggestedcitationforthischapterofnotesis: Schwarz,C.J.(2015). TwoFactorDesigns-Single-sizedExperimentalunits-CRandRCB designs. InCourseNotesforBeginningandIntermediateStatistics. Availableathttp://www.stat.sfu.ca/~cschwarz/CourseNotes. Retrieved2015-08-20. 10.1 Introduction So far we’ve looked at two different experimental designs, the single-factor completely randomized design(1-factorCRD),andthesingle-factorrandomizedcompeteblockdesign(1-factorRCB). Bothdesignsinvestigatedifdifferencesinthemeanresponsecouldbeattributedtodifferentlevels ofasinglefactor. However, inmanyexperiments, interestliesnotonlyintheeffectofasinglefactor, butinthejointeffectsof2ormorefactors. Forexample: • Yieldofwheat. Theyieldofwheatdependsuponmanyfactors-twoofwhichmaybethevariety andtheamountoffertilizerapplied. Thishastwofactors-(1)varietywhichmayhavethreelevels representingthreepopulartypesofseeds,and(2)theamountoffertilizerwhichmaybesetattwo levels. • Pesticidelevels. Thepesticidelevelsmaybemeasuredinbirdswhichmaydependuponsex(two levels)anddistanceofthewinteringgroundsfromagriculturalfields(threelevels). • Performanceofaproduct. Thestrengthofpapermaydependupontheamountofwateradded (twolevels)andthetypeofwoodfiberusedinthemix(threelevels). 527 (cid:13)c2015CarlJamesSchwarz 2015-08-20 CHAPTER10. TWOFACTORDESIGNS-SINGLE-SIZEDEXPERIMENTALUNITS-CRAND RCBDESIGNS There are many ways to design experiments with multiple factors - we will examine three of the mostcommondesignsusedinecologicalresearch-thecompletelyrandomizeddesign(thischapter),the randomizedblockdesign(thischapter),andthesplit-plotdesign(nextchapter). Asnotedmanytimesinthiscourse,itisimportanttomatchtheanalysisofthedatawiththewaythe datawascollected. Beforeattemptingtoanalyzeanyexperiment,thefeaturesoftheexperimentshould beexaminedcarefully. Inparticular,caremustbetakentoexamine • thetreatmentstructure; • theexperimentalunitstructure; • therandomizationstructures; • thepresenceorabsenceofbalance; • ifthelevelsoffactorsarefixedorrandomeffects;and • theassumptionsimplicitlymadeforthedesign. Ifthesefeaturesarenotidentifiedproperly,thenanincorrectdesignandanalysisofanexperimentwill bemade. 10.1.1 Treatmentstructure Thetreatmentstructurereferstohowthevariouslevelsofthefactorsarecombinedintheexperiment. The first step in any design or analysis is to start by identifying the factors in the experiment,their associatedlevels,andthetreatmentsintheexperiment. Treatmentsarethecombinationsoffactorlevels thatare‘applied’1toexperimentalunits. Thetwo-factordesignhas,asthenameimplies,twofactors. WegenericallycalltheseFactorAand FactorBwithaandblevelsrespectively. Wewillexamineonlyfactorialtreatmentstructures,i.e. every treatmentcombinationappearssomewhereintheexperiment. Forexample,ifFactorAhas2levels,and FactorBhas3levels,thenall6treatmentcombinationsappearintheexperiment. Whyfactorialdesigns? Why do we insist on factorial treatment structures? There is a temptation to investigate multi-factor effects using a ‘change-one-at-time’ structure. For example, suppose you are investigating the effects of process temperature (at two levels, H & L), fiber type (at two levels - deciduous and coniferous) and initial pulping method (at two levels - mechanical or chemical) upon the strength of paper. In the ‘change-one-at-a-time’treatmentstructure,thefollowingtreatmentcombinationswouldbetested: 1. L deciduous mechanical 2. H deciduous mechanical 3. L coniferous mechanical 4. L deciduous chemical 1Recallthatinanalyticalsurveys,thefactorlevelscannotbeassignedtounits(e.g. youcan’tassignsextoananimal)andso thekeypointisthatunitsarerandomlyselectedfromtherelevantpopulation. 528 (cid:13)c2015CarlJamesSchwarz 2015-08-20 CHAPTER10. TWOFACTORDESIGNS-SINGLE-SIZEDEXPERIMENTALUNITS-CRAND RCBDESIGNS The researcher then argues that the effect of fiber type could be found by examining the difference in strengthbetweentreatments(1)and(3);theeffectofpulpingmethodcouldbefoundbyexaminingthe difference in strength between treatments (4) and (1); and the effect of process temperature could be foundbyexaminingthedifferenceinstrengthbetweentreatments(1)and(2). Thisisvalidprovidedthattheresearcheriswillingtoassumethetreatmenteffectsareadditive, i.e.,thattheeffectofprocesstemperatureisthesameatalllevelsoftheotherfactors;thattheeffectof fibertypeisthesameatalllevelsoftheotherfactors;andthattheeffectofinitialpulpingmethodisthe sameatalllevelsoftheotherfactors. Unfortunately,thereisnomethodavailabletotestthisassumption withthesetoftreatmentslistedabove. Itisusuallynotagoodideatomakethisverystrongassumption-whathappensiftheassumptionis nottrue? Inthepreviousexample,itmeansthatyour‘effects’areonlyvalidfortheparticularlevelsof theotherfactorsthathappenedtobepresentinthecomparison. Forexample, theprocesstemperature effectwouldonlybevalidfordeciduousfibersourcesthataremechanicallypulped. Asuperiortreatmentstructureisthefactorialtreatmentstructure.Inthefactorialtreatmentstructure, every combination of levels appears in the experiment. For example, referring back to the previous experiment,allofthefollowingtreatmentswouldappearintheexperiment: 1. L deciduous mechanical 2. H deciduous mechanical 3. L coniferous mechanical 4. H coniferous mechanical 5. L deciduous chemical 6. H deciduous chemical 7. L coniferous chemical 8. H coniferous chemical Now,themaineffectsofeachfactorarefoundas: • maineffectoftemperature-treatments1,3,5,7vs.2,4,6,8 • maineffectofsource-treatments1,2,5,6vs.3,4,7and8 • maineffectofmethod-treatments1,2,3,4vs.5,6,7,8 Eachmaineffectwouldbeinterpretedatthe‘averagechange’overthelevelsoftheotherfactors. Inaddition,itispossibletoinvestigateifinteractionsexistbetweenthevariousfactors.Forexample, istheeffectofprocesstemperaturethesameformechanicalandchemicalpulpingmethods? Thiswould beexaminedbycomparingthechangein(1)+(3)vs.(2)+(4)[representingtheeffectoftemperaturefor mechanicallypulpedwood]andthechangein(5)+(7)vs.(6)+(8)[representingtheeffectoftemperature for chemically pulped wood]. Can you specify how you would investigated the interaction between temperatureandsource? Whataboutbetweensourceandmethodofpulping? Alloftheseareknownas twofactorinteractions. Theconceptofatwo-factorinteractioncanalsobegeneralizedtothree-factorandhigherinteraction termsinmuchthesameway. 529 (cid:13)c2015CarlJamesSchwarz 2015-08-20 CHAPTER10. TWOFACTORDESIGNS-SINGLE-SIZEDEXPERIMENTALUNITS-CRAND RCBDESIGNS Whynotfactorialdesigns? While a factorial treatment structure provides the maximal amount of information about the effects of factors and their interactions, there are some disadvantages. In general, the number of treatments that willappearintheexperimentisequaltotheproductofthelevelsfromallofthefactors.Inanexperiment withmanyfactors,thiscanbeenormous. Forexample,ina10factordesign,witheachfactorat2levels, thereare1024treatmentcombinations. Itturnsoutthatinsuchlargeexperiment,therearebetterways toproceedthatarebeyondthescopeofthiscourse-anexampleofwhichisafractionalfactorialdesign which selects a subset of the possible treatments to run with the understanding that the subset chosen losesinformationonsomeofthehigherorderinteractions.Ifyouarecontemplatingsuchanexperiment, pleaseseekcompetenthelp. As well, in some cases, interest lies in estimating a response surface, e.g. factors are continuous variables(suchatemperature)andtheexperimenterisinterestedinfindingtheoptimalconditions. This gives rise to a class of designs called response surface designs which are beyond the scope of this course. Again,seekcompetenthelp. Displayingandinterpretingtreatmenteffects-profileplots An important part of the design and analysis of experiment lies in predicting the type of response ex- pected - in particular, what do you expect for the size of the main effects and do you expect to see an interaction. During the design phase, these are useful to determining the power and needed sample sizes for an experiment. Duringthe analysis phase, these valuesand plots helpin interpreting theresults of the statisticalanalysis. Withtwofactors(AandB)eachattwolevels,youcanconstructaprofileplot. Theseprofileplots showtheapproximateeffectofbothfactorssimultaneously. Thekeythingtolookforisthe‘parallelism’ofthetwolines. Profileplotswithnointeractionbetweenfactors Forexample,considerthetheoretical[itistheoreticalbecauseitshowsthepopulationmeanswhichare neverknownexactly]profileplotofthemeanresponsesbelow: 530 (cid:13)c2015CarlJamesSchwarz 2015-08-20 CHAPTER10. TWOFACTORDESIGNS-SINGLE-SIZEDEXPERIMENTALUNITS-CRAND RCBDESIGNS Inthisplot,theverticaldistancebetweenthetwoparallellinesegmentsistheeffectofFactorB,i.e., whathappenstothemeanresponsewhenyouchangethelevelofFactorB,butkeepthelevelofFactor Aconstant. ThemaineffectofFactorBistheAVERAGEverticaldistancebetweenthetwolineswhen averaged over all levels of Factor A. Notice that if the lines are parallel, the vertical distance between thetwolinesisconstant-thisimpliesthattheeffectofFactorB(theverticaldistancebetweenthetwo lines) is the same regardless of the level of Factor A and the effect of Factor B and the main effect of FactorBaresynonymous. Inthiscase,wesaythatthereisNOINTERACTIONbetweenFactorAand FactorB.Similarly, theeffectofFactorAisthechangeinthelinebetweenthetwolevelsofFactorA ataparticularvalueofFactorB,i.e.,theverticalchangeineacheachlinesegment. Themaineffectof FactorAistheAVERAGEchangewhenaveragedoveralllevelsofFactorB.Noticethatifthelinesare parallel,theverticalchangeisthesameforbothlines-thisimpliesthattheeffectofFactorAisthesame regardlessofthelevelofFactorBandthattheeffectofFactorAissynonymouswiththemaineffectof FactorA.Onceagain,thereisnointeractionbetweenAandB. Profileplotswithinteractionbetweenfactors Nowconsiderthefollowingtheoreticalprofileplot: 531 (cid:13)c2015CarlJamesSchwarz 2015-08-20 CHAPTER10. TWOFACTORDESIGNS-SINGLE-SIZEDEXPERIMENTALUNITS-CRAND RCBDESIGNS Inthisplot,theverticaldistancebetweenthelinesegmentsCHANGESdependingonwhereyouare inFactorA.ThisimpliesthattheeffectofFactorBchangesdependinguponthelevelofA,i.e.,there is INTERACTION between Factor A and B. The main effect of Factor A is the average effect when averaged over levels of B. In this case the main effect is not very interpretable (as will be seen in the plotsbelow). Similarly, theverticalchangeforeachlinesegmentisdifferentforeachsegment-again the effect of Factor A changes depending upon the level of Factor B - once again there is interaction betweenAandB. Theplotsfromanactualexperimentmustbeinterpretedwithagrainofsaltbecauseeveniftherewas nointeraction,thelinesmaynotbeexactlyparallelbecauseofsamplingvariationsinthesamplemeans. Thekeythingtolookforisthedegreeofparallelism. Anditdoesn’tmatterwhichfactorisplottedalong thebottom-theplotsmaylookdifferent,butyouwillcometothesameconclusions. Ifthereisinteraction,thelinesegmentsmayevencrossratherthanremainingseparate. Illustrationsofvarioustheoreticalprofileplots 532 (cid:13)c2015CarlJamesSchwarz 2015-08-20 CHAPTER10. TWOFACTORDESIGNS-SINGLE-SIZEDEXPERIMENTALUNITS-CRAND RCBDESIGNS • NomaineffectofFactorA(averageoflinesisflat);smallmaineffectofFactorB(iftherewasno maineffectofFactorBthelineswouldcoincide);andnointeractionofFactorsAandB. • LargemaineffectofFactorA;smallmaineffectofFactorB(averagedifferencebetweenlinesis small);andnointeractionbetweenFactorsAandB. • NomaineffectofFactorA;largemaineffectofFactorB;andnointeractionbetweenFactorsA andB. 533 (cid:13)c2015CarlJamesSchwarz 2015-08-20 CHAPTER10. TWOFACTORDESIGNS-SINGLE-SIZEDEXPERIMENTALUNITS-CRAND RCBDESIGNS • LargemaineffectofFactorA;largemaineffectofFactorB;andnointeractionbetweenFactors AandB. • NomaineffectofFactorA;nomaineffectofFactorB;butlargeinteractionbetweenFactorsA and B. This illustrates the dangers of investigating ‘main effects’ in the presence of interaction (why? -agoodexamquestion!). • LargemaineffectofFactorA;nomaineffectoffactorB:slightinteraction. Again,thisdiagram illustratesthefollyofdiscussingmaineffectsinthepresenceofaninteraction(why?). 534 (cid:13)c2015CarlJamesSchwarz 2015-08-20 CHAPTER10. TWOFACTORDESIGNS-SINGLE-SIZEDEXPERIMENTALUNITS-CRAND RCBDESIGNS • NomaineffectofFactorA;largemaineffectofFactorB;largeinteractionbetweenFactorAand B.Asbefore,theremaybeproblemsininterpretingmaineffectsinthepresenceofaninteraction (why?). 535 (cid:13)c2015CarlJamesSchwarz 2015-08-20
Description: