RDP4软件分析指南

重组检测软件RDP4的应用汇报人：赵烨IntroductionRDP4(RecombinationDetectionProgramversion4)是一个检测和分析重组信号的软件，包括7种基本检测程序（RDPmethod,GENECONV,Bootscanning,MaxChi,Cimaera,3SEQandSiScan）和4种附加检测程序（LARD,PHYLPRO,distanceplotsandTOPAL）。操作基本步骤•打开一个.meg格式的文件：左击鼠标open按钮。•点击页面顶端options按钮，进入General页面选择一系列参数：•左击主界面上的X-over按钮开始进行重组分析。•分析完毕后出现四个界面，顺时针方向依次为：•Sequencedisplay•Therecombinationinformationdisplay•Schematicsequencedisplay•PlotdisplayRecordsignificantevidenceofrecombinationMoveaslidingwindowacrosssub-sequenceandcalculatepairwiseidentitiesMultiplesequencealignmentSelectthreesequencesanddiscardallnon-informativesitesInformation-richsub-sequenceCheckforevidenceofrecombinationCalculatesignificancewhere:GisthetotalnumberofofpossiblesequencetripletsListheLengthofthesequenceNisthelengthoftheputativelyrecombinantregionmistheproportionofnucleotidesincommonbetweentheputativerecombinantandparentalsequencesintherecombiantregionpistheproportionofnucleotidesincommonbetweentheputativerecombinantandparentalsequencesintheentiresequence.Repeatwiththenextthreesequences()P=N!m!(N-m)!pNX(1-p)N-mXLNΣm=MNGXAACGCGATTGCAGGAAGGCATATGTTATGGCATAAGGCGATAGCAGGTGGCCTTACATTATGGCATAAGGCGATTCCTGGAAGCCTTACGTAATGGCATAAGGTGATAGCAGGTAGCCTTACATAATCGCATAAGGCGATAGCAGGTAGGCTTACGTTATCGCATAAGGCGATAGCAGGTAGGCTTATATTACGGCATGTCTACTCGATCTGAAGATGTTGAGATGTTATCA1760152122823043Positioninalignment1.00.50.0Pairwiseidentity1.07X101.07X10--66PositionsofinformativesitesPotentialrecombinantregionMajorparent:recombinantplotMinorparent:recombinantplotMinorparent:majorparentplotP-Value1760152122823043Positioninalignment1.00.50.01760152122823043Positioninalignment1.00.50.0Pairwiseidentity1.07X101.07X10--66PositionsofinformativesitesPotentialrecombinantregionMajorparent:recombinantplotMinorparent:recombinantplotMinorparent:majorparentplotP-ValueFigure8.TheoriginalRDPmethod.ATheanalysisprocedure.BAnexamplepairwiseidentityplot.BRecordsignificantevidenceofrecombinationMoveaslidingwindowacrosssub-sequenceandcalculatepairwiseidentitiesMultiplesequencealignmentSelectthreesequencesanddiscardallnon-informativesitesInformation-richsub-sequenceCheckforevidenceofrecombinationCalculatesignificancewhere:GisthetotalnumberofofpossiblesequencetripletsListheLengthofthesequenceNisthelengthoftheputativelyrecombinantregionmistheproportionofnucleotidesincommonbetweentheputativerecombinantandparentalsequencesintherecombiantregionpistheproportionofnucleotidesincommonbetweentheputativerecombinantandparentalsequencesintheentiresequence.Repeatwiththenextthreesequences()P=N!m!(N-m)!pNX(1-p)N-mXLNΣm=MNGXAACGCGATTGCAGGAAGGCATATGTTATGGCATAAGGCGATAGCAGGTGGCCTTACATTATGGCATAAGGCGATTCCTGGAAGCCTTACGTAATGGCATAAGGTGATAGCAGGTAGCCTTACATAATCGCATAAGGCGATAGCAGGTAGGCTTACGTTATCGCATAAGGCGATAGCAGGTAGGCTTATATTACGGCATAACGCGATTGCAGGAAGGCATATGTTATGGCATAAGGCGATAGCAGGTGGCCTTACATTATGGCATAAGGCGATTCCTGGAAGCCTTACGTAATGGCATAAGGTGATAGCAGGTAGCCTTACATAATCGCATAAGGCGATAGCAGGTAGGCTTACGTTATCGCATAAGGCGATAGCAGGTAGGCTTATATTACGGCATGTCTACTCGATCTGAAGATGTTGAGATGTTATCGTCTACTCGATCTGAAGATGTTGAGATGTTATCA1760152122823043Positioninalignment1.00.50.0Pairwiseidentity1.07X101.07X10--66PositionsofinformativesitesPotentialrecombinantregionMajorparent:recombinantplotMinorparent:recombinantplotMinorparent:majorparentplotP-Value1760152122823043Positioninalignment1.00.50.01760152122823043Positioninalignment1.00.50.0Pairwiseidentity1.07X101.07X10--66PositionsofinformativesitesPotentialrecombinantregionMajorparent:recombinantplotMinorparent:recombinantplotMinorparent:majorparentplotP-ValueFigure8.TheoriginalRDPmethod.ATheanalysisprocedure.BAnexamplepairwiseidentityplot.BFigure1.ThemaincomponentsoftheRDPinterface.Commandbuttons命令按钮Treedisplay进化树显示Matrixdisplay矩阵显示Recombinationinfo.Display重组信息显示Sequencedisplay序列显示Plotdisplay绘图显示Schematicsequencedisplay图解序列显示Identitydisplay同源性显示RDP4界面的主要组成部分分析进度条Positionindicator位点指示Identitydisplay同源性显示Sequencedisplay序列显示SequencenamesTogglesequencedisplayIdentityindicatorToggleidentitydisplaysequencedisplay.左击序列中的部位可以显示不同颜色代表的含义鼠标悬空在某个核苷酸上会显示出该核苷酸处于何序列的具体位置右击鼠标可以保存不同形式你想要的序列Warningsandadditionalmethodspecificinfo.P-ValuesProbablerecombinant(recombinant)andparentalsequencesBreakpointpositions:Insequence(inalignment)MethodusedtodetectrecombinantregionTherecombinationinformationdisplay现以下情况，该界面还会出现warning标示（红色）：（1）在比对序列中只有一个可能为父母代毒株的序列（2）有可能(约30%或更大的可能)误认为重组序列(即实际上父母代的序列中的某个才是真正的重组毒株)如果是这样会在后面显示出实际上可能为重组毒株的父母代毒株的名字。（3）无法识别出一个或全部两个突变位点。（4）一个或两个突变位点是错位的。（5）重组信号微弱（6）如果复合信号可能是一个分析错误的人工制品。“confirmationtable”部分表明了用不同方法检测出发生该重组事件的毒株数和关于目前检测到的重组事件的符合程度Confirmationtable下面是一个总结性的柱状图，对于99%的用户来说前三条柱状图代表的是有用的信息。柱状图下面的分数大于60%代表这该毒株几乎确定为重组毒株。大于40%小于60%的分数代表软件可能犯了错误，但也可能没有。小于40%表示该毒株很可能不是重组毒株。TheschematicsequencedisplayCurrentviewCyclethroughdisplayoptions改变视图颜色Recombinantregion重组区域Backgroundsequence背景序列主要亲本与重组序列的同源性潜在的重组区域P-value次要亲本与重组序列的同源性主要亲本与次要亲本的同源性ThePlotDisplay“Recombinant”sequence“Parental”sequencesPortionofalignmentusedtodrawtreeCyclethroughtreesViewalltreesSimultaneouslySwitchtorecombinationinformationdisplaySwitchtocompatibilitymatrixdisplayFigure6.Thedend