primer5设计pcr引物的步骤

《primer5设计pcr引物的步骤》由会员上传分享，免费在线阅读，更多相关内容在工程资料-天天文库。

1、Primer5设计引物打开软件/Pri>erPre>ierFurictionHelpINew►DNASequence1Open»groteinSequencePreferencesDegenerat©basesCtrl+RQuit1C:DocumgntsandSettingsAdministrator桌面123.seq2C：PROGRA"1PRIMER"1dataDemoTNF・ALN复制基因序列GTGAAACGCTTTTTTCTGGTGGCGATCGCCTCTGTCCTAnTTTCTTTAACACCA

2、TGGTclGGCAGTGCCAATGCGGTGGAGTT^CCGAAAGCACCCGCACCATTCCCCTCGATG.AAGCCGGTGGCACCACAACCTTAACCGCCCGCCAGTTCACCAACGGCCAAAAGATTTTTGIJggacacctgcaccca.atgtcacctccagggtaaaacc^4aactaata.aiaacgttagt

3、TTAGGCTTGGCAGACTTGGCTGGTGCCGAGCCCCGTCGGGACAACGTCCTGGCCTTGdTGGAATTTCTCAAGAATCCC

4、AAGAGTTATGACGGTGAAGATGACTATTCGGAATTACAljCCCAATArTTCCCGTCCCGACATCTACCCAGAAATGAGAAACTACACCGAAGACGACAlTCTTTGATGTGGCTGGTIACACCCTCATTGCTCCCAAATTAGATGAACGGTGGGGTGG^IACCATCTACTTCTAG^-'粘贴选项ASis-OK点击primer按钮，弹出菜单后点击search按钮61QGCAGTGC121GGTGGCACGeneTank一NevSequence5,SeqNo

5、

6、

7、冋仙

8、

9、Find

10、〔FindNext

11、目因［dsDNA11BFunction:Translation:181241301361TCCCGTCC421GCTGGTTJ481TAGACCTGCACTTGGCAG；CTCAAGAJ选择PCRprimer,Pairs参数，并选择所需PCR产物长度，OKFunction:GGCAGTGC181ACCTGCA：rTTQGCAG；241301CTCAAGA210SenseAnti-sense68.848.0Translation:ActiveSequence:[NewSequen

12、ceTranslations:能nzymeMotif5・SeqNo1213€1421431SearchFor:了PCRPrimers^)SequencingPrimersGGTGGCAC.一―pnmen(^DTCCCGTCCGCTGGTT；TAGDirectSelectSearchType:SensePrimerGTGAAAUGLT-TTTr-TQQTccr^ZSSQSAnti>sensePrimerCompatiblewithSensePrimerCompatiblewithAnti-sensePrimerSearc

13、hRanges:PrimerLength:Rating3-CACTTTGCGAAA.IlllllllllllST^AAACCXCTTTHairpinDimerFalsePrimingCrossDimerMostStableHairpin:,Gs-2.9Pccal$/mol](3*Hairpin)0Sense

14、FoundFound

15、None

16、Found

17、rTTTTTTCGCAAAGTG5*LrTGCTGGrGA3•Anti-senseFounci

18、Found

19、None

20、"I1►Product会弹岀搜索结果菜单，点击OKS

21、earchProgressPrimerSearchResults:TotalPossible1398

22、B

23、Tm438凰GC%91

24、H

25、Degeneracy0[H]3'EndStability219

26、回

27、GCClamp40

28、画

29、Redundancy457[fl]Repeats/Runs27回Dimer/Hairpin60[□1FalsePriming[E]OptimalPrimers66RemainingRejectedSense:Anti13984389102035546S196757：✓ok]^CancelStrin

30、gency:[H]VeryHigh凰High回P»1oderate回Low回VeryLow回ManualHPairsFound85PrimerPairs:在搜索出的结果列表里选择TM合适、GC含量高、无各种BUG的引物，点击editprimer,将所得引物复制RatingSeqNoLengthTm[°C]GC%AG[kcal5•m