Quick Start Guide - Thermo Fisher Scientific
Contents
1. Citenerabescrption deste TGA Find ORE x KA i standard Fields Apa li pean nd PTT Silent Mutations b K Annotations Eze HE era _ J Restriction Methylation slap Translation gt e Endre products rebated to dpt Axel Chan r Edea Aaa Restriction Analyses gt a5 dt F Back Translation MA ap BioAnnotator b 3 C3DirectiForwerd strand S Z el z muet u Lis F li ConomBanch Tool E Steading Frane 1 B ORE apa b m IL af er Lede P Tanen trie t QR D PENAS 2 N EmB n PEST vera Run Analysis Monitor Reading Frame 3 74 ORFs ES 4 u quM Cede Beverse xd rond 1 Carseat Web Analyses B ators rl ai and i Cherating Frame 5 RF L o i Steading Frame 2 7 ftFs P S F v ae Miellit i heading Frame 3 26 ORFs x a i Ht Eam BE 2445 ORF Setup Tw T yo b l Heat lagas X i Erma oid wh Start Codons ATG GTG PEU TNT Emel aa 00 X Aral 300 a i Araligi Tie Smal shag StopCodons TAA TGA TAG Cancel Kral aac Apel Bee aurea Faits Default Start amp Stop L Include Stop Codon in ORF E 1401 DRF Types pM IV Complete ORFs which have both start and stop E codons defined as above Minimum Size 150 bp M Nested C guum ORFs which do not have both defined s MRTA G AGAAACGTAA AATGATATAA ATATCAA Incomplete start or stop codons i i i EERCTTOC TETTIGCATT TTACTATATT TATAOTT Kiko Si bp 1601 ATTAAATTAG ATTTTOCATA AAAAACAGAT TACATAATAC TGTAAAACAC AA2. i Gateway Sites 0 3 terminus EI Chess wine ntn User Defined ze VectorNTI DNA RNA Molecule From document seq Ext AX Ei File Edit View Primer Design Analyses Cloning Gel Analysis List Window Align Assemble Tools Help x 4 d 2 AROSA il B CAAS TE PE dzaT iadd C General Description Bam HT a33 daaT ag3 3 C Standard Fields Peas Pes PTUS McoT gza ApaLlliars A Comments Y 1 O LJ Annotations o C Feature Map 5 C Restriction Methylation Map seq nazop I TGCCCAAGA AGAAGCCCAC CCCCATCCAG CTGAACCCCG CCCCCCCCGA CGGCAGCGCC GTGAACGGCA CCAGCAGCGC CGAGACCAL ACGGGTTCT TCTTCGGGTG GGGGTAGGTC GACTTGGGGC GGGGGGGGCT GCCGTCGCGG CACTTGCCGT GGTCGTCGCG GCTCTGGTT Pstl LAAN 101 TGCAGAAGAA GCTGGAGGAG CTGGAGCTGG ACGAGCAGCA GCGGAAGCGG CTGGAGGCCT TCCTGACCCA GAAGCAGAAG GTGGGCGAG ACGTCTTCTT CGACCTCCTC GaccTcGacc TGCTCGTCGT CGCCTTCGCC GACCTCCGGA AGGACTGGGT CTTCGTCTTC CACCCGCTC 201 CGACTTCGAG AAGATCAGCG AGCTGGGCGC CGGCAACGGC GGCGTGGTGT TCAAGGTGAG CCACAAGCCC AGCGGCCTGG TGATGGCCCt GCTGAAGCTC TTCTAGTCGC TCGACCCGCG GCCGTTGCCG CCGCACCACA AGTTCCACTC GGTGTTCGGG TCGCCGGACC ACTACCGGGt Pstl SLOD 301 CACCTGGAGA TCAAGCCCGC CATCCGGAAC CAGATCATCC GGGAGCTGCA GGTGCTGCAC GAGTGCAACA GCCCCTACAT CGTGGGCTT GTGGACCTCT AGTTCGGGCG GTAGGCCTTG GTCTAGTAGG CCCTCGACGT CCACGACGTG CTCACGTTGT CGGGGATGTA GCACCCGAL 4 liil b Ready i bp E Figure 11 In Silico Gene Synthesis with ReGENerator 12 Identifying Open
3. ALF file Print Ctrl P From EMBL file Print Preview From Text file Print setup pe From ESD file Exit From Phred files From Phrap ACE file a Eontigexpress Project Untitled S ze Folder Of Fragments Project Edit View Assemble Align Analyses Tools Help History View 12 fragment s in project B Fragments MAIN Name Length Original length zu ONE17KANR 759 759 za OMEZKANR 747 747 zu ONESKANR 755 755 zu ONESKANR 756 756 za ONE6KANR 770 770 ui 4 z 5 mq a c w Ed 1 CAATT NTCGA TGATG GTTGA GATGT GTATA AGAGA 51 TCCTG CATTA GGAAG CAGCC CAGTA GTAGG TTGAG 101 CGCCG CAAGG AATGG TGCAT GCAAG GAGAT GGCTG 151 GCCAC GGGGC CTGCC ACCAT ACCCA CNGCC GAAAC 3 7 ONE17KANR 759 amp General Description Switch to info paneOText pane Creation Date 12 11 PM November 2 Last Modification Date 12 11 PM Nove Cy Chromatogram Data 201 CGAAG TGGCG AGCCC GATCT TCCCC ATCGG TGATG ex ONEBKANR 764 764 Start date of the gel run 05 42 PM Ja 251 CCAGC AACCG CACCT GTGGC GCCGG TGATG CCGGC ONESKANR 758 758 Stop date of the gel run 09 57 PM Jar 301 GT G GGATC TGGCT AGCGA TGACC CTGCT GATTG za ONELOKANF 758 758 Scaling Factors For the average signals 351 NCGGG TGCGG GACGG CGTTA CCAGA AACTC AGAAG ONELIKANR 755 755 za OMET3KANR 819 819 ONE15KANF 784 784 za ONET6KANF 767 767 401 ccGac TCTG CGGCA GTTTA CGAGA GAGAT GATAG are maar seme oss s
4. Be Expressed window and inserting deleting or replacing single or multiple amino acids e Click on the Change button to select the desired Codon Usage Table for the organism e Select the desired Genetic Code from the dropdown menu e Add specific binding sites or adaptors to the ends of the DNA sequence by selecting the desired terminus 5 or 3 and clicking on the appropriate button e g Restriction Sites Gateway Sites or User Defined To view the back translated DNA click View In Silico DNA To send the DNA sequence to GENEART for synthesis click Send for Synthesis Protein to be Expressed 1 MPKKEPTPIO LNPAPI VNGTSSAETN LEALOKKL 51 LEAFLTOKOK VGELKDDDFE KISELGAGNG GVVFEVSH 101 HLEIKPAIRN OIIRELOVLH ECNSPYIVGF YGAFYSDG EERE 151 SLDOVLEKAG RIPEQILGEV SIAVIKGLTY LREKHEIM i 201 SRGEIKLCDF GVSGOLIDSM ANSFVGTRSY MSPERLOG ta 1 nii v 251 GLSLVEMAVG RYPIPPPDAK ELELMFGCQV EGDAAETP ai e zi To Make Mutations gt JE n Add Attachments to In Silico DMA Sequence Substitutions Highlight the amino acid s and type in your desired changes AJ 7 m Deletions Highlight the amino acid s and use the Delete or Backspace key Mi ni ae ThE Note Deletions will not be marked in bs Protien Sequence Pane MA Hi ABA 27 Attach to Choose Attachment Type Insertions Place your cursor upstream of the insertion point and type in your desired 1 MG i ib 15 IL MIB LE ce B terminus Restriction Sites Leid GEL EL Ba H UT JEN GU
5. Utilities Genomic Data Analysis Licensing and Support e Invitrogen by Life technologies Figure 1 QuickStart Page You can configure the software to open both the Molecule Viewer and Vector NTI Explorer when you select Vector NTI from the Start menu 1 In the Molecule Viewer window go to the View menu and select Options 2 In the General tab of the dialog select the Open Local Database at Startup checkbox 3 Click OK to make the change Local Database Vector NTI Explorer is the main tool for accessing the information in your local Vector NTI Advance database Using the Explorer you can import open export and organize molecules and other database items and launch other Vector NTI Advance modules Figure 2 To launch Vector NTI Explorer e On QuickStart Page click on Launch Local Database e Inthe Molecule Viewer click on the Local Database icon hey e From the Windows Start menu select Programs gt Invitrogen gt Vector NTI Advance 11 gt Vector NTI Explorer e The local database in Vector NTI Advance contains records for different types of molecular biology objects Each database record includes all the information for that object e g a DNA molecule record includes the DNA sequence defined features of the molecule and other information Objects in the database can include molecules analysis results BLAST search results citations and other types of information Import create m
6. 1 0 Dacuments 5 Web Beta 7 WebEx El T My Computer E E 4 My Network Places Year Make New Folder Cancel o gt a E r 2008 Set Reminder Exit Database Backup e From Vector NTI Explorer menu select Database gt Database Restore Database A Search Change Author Date Column Fields Manager Frequent Contact Manager Database Backup b Database Restore R Database Cleanup e Local Shared Data Exchange Select Local Database Figure 4 Choose Badu Dinectony kl LET S Updaber i3 updates a Ucer Training t 9 C23 VMTE Sugg 4 Eo vMIE w nson 11 0 Documents gt Web Beta H p web x Ej XD Mi Computer si See ee al unde 705i Database Restore Molecule Viewer The Molecule Viewer displays information about DNA RNA and protein molecules To launch the Molecule Viewer e Click on Launch Molecule Viewer on the QuickStart page or e From the Windows Start menu select Programs gt Invitrogen gt Vector NTI Advance 11 gt Vector NTL or e Double click on a molecule name in the Vector NTI Explorer To open a molecule from within the Molecule Viewer click on the Open button toolbar and select the molecule name from the dialog box on the main The molecule will be loaded into the Molecule Viewer 3 Vector AT rutorial 1 zB Ei File Edit View Primer Design Analyses Cloning GelAnalysis List Window Align Asse
7. 1364 bp Us Figure 5 Molecule Viewer window for a DNA molecule The Molecule Viewer window has different panes for displaying different types of information about the molecule as shown in Figure 5 Click inside a pane to make it the active pane The available tools and right click menu options will change depending on which pane is active Use tools on the dropdown menus and toolbars to add information about the molecule and perform various analysis functions as described in the step by step instructions on the following pages Multiple molecules can be displayed in separate windows of the Molecule Viewer Selecting and Editing Molecule Sequences In the Molecule Viewer you can select part of a molecule sequence in several different ways e Hold down the mouse button and drag the cursor across the sequence in the Sequence Pane or Graphics Pane Figure 6 e Go to the Edit menu select Set Selection and enter the sequence base pair range in the dialog box e Click on a defined feature in the Graphics Pane ATG e Click on a defined feature in the Text Pane and click on Find ra on the main toolbar The selected sequence will appear highlighted in both the Graphics Pane and the Sequence Pane S usu FLD1 promoter a ia GCATGCAGGA ATCTCTGGCA CGGTGCTAAT GGTAGTTATC CAACGGAGCT GAGGTAGTCG ATATATCTGG ATATGCCGCC TATAGGATA CGTACGTCCT TAGAGACCGT GCCACGATTA CCATCAATAG GTTGCCTCGA CTCCATCAGC TATATAGACC TATACGGCGG ATATCCTAT GGGT
8. 3 2464 Bg E Complexity and similarity plots H EJ e El e Absolute Complexi ill pSP71 0 0012 z i solute pSP70 s ir a rie Relatedness pSP72 0 0046 tre e Sequence alignment 1 LJ Consensus Figure 14 Aligning molecules 15 Contig Assembly Vector NTI Advance 11 5 can be used to assemble DNA fragments both text sequences and chromatograms from automated sequencers into longer contiguous sequences or contigs The tool for doing this is called ContigExpress In Vector NTI Explorer or the Molecule Viewer 1 2 10 16 Go to the Assemble menu and select ContigExpress Open New Assembly Project Figure 154 In the ContigExpress Project Explorer go to the Project menu and select Add Fragments gt Select your fragment file type from the submenu list The Import Sequence dialog will open In the Import Sequence dialog navigate to the directory containing your fragment sequence files Select the files and click on Open Depending on the file type you may be prompted to list the fragments by their Windows file names or by their internal fragment names Select the desired option The fragments will be loaded in the ContigExpress Project Explorer To view a particular fragment double click on it in the Project Explorer list It will be loaded into the Fragment Viewer When you are ready to perfo
9. C NVNTI Database Database Backup Restore It is strongly recommended that the local database be backed up routinely You may launch the Database Backup manually or use the Database Backup Reminder to trigger the task automatically as configured Figure 3 To manually perform Database Backup e From the Vector NTI Explorer menu select Database gt Backup Database Now To configure the Database Backup Reminder e From the Vector NTI Explorer menu select Database gt Database Backup Reminder e To set a specific date or interval e g backup every 15 days click on Set Reminder Database A Search Change Author Date Column Fields Manager Frequent Contact Manager Database Backup Database Backup Reminder X TR beken Restos Backup Database Now Database Cleanup e Local Shared Data Exchange Backup Reminder Conhautation Select Local Database below Set Date Remind on a Date Date Month Set Number of Days Switch Reminder Off Reminder OFF Figure 3 To restore a database You can store manage and Find critical sequence and the other molecular biology data in the Vector NTI Advance Local Database Please remember to backup your data regularly You can configure through the backup reminder Choose Backup Directory i Updaters El ic User Training I Vi 3 IACKLIP FOLDER i7 VMTI Support t VMTI Version 1
10. CATATCCA GTCACTATGG COGEEGCATT AGGCACCCCA GECTTTA TAATTTAATG TALAACGTAT TTTTTGTCTG ATGTATTATG LCATTTTGTG TTGTATAGGT CAGTGATACC GCCOGCGTAA TCC TOGGGT COGAAAT Undefined Pd al Insae m bp et bo 543 bo 1022 bes Figure 12 Identifying ORFs 13 Creating a Restriction Map Vector NTI Advance 11 5 can analyze a DNA RNA molecule and identify the restriction sites in it using the software s comprehensive library of restriction enzymes With a DNA or RNA molecule open in the Molecule Viewer 1 Go to the Analyses menu and select Restriction Analyses gt Restriction Sites Figure 13 2 In the Restriction Map Setup dialog review the list of restriction enzymes in the Use Enzymes field These are the enzymes that will be used to identify the restriction sites Click on the lt Add gt Remove and gt gt Remove All buttons to add and remove enzymes from the list Note If you click on lt Add the Choose Database Enzymes dialog will open listing all the enzymes in the database Select enzymes in the list by clicking on them or click on the Select All button and then click on the OK button to add them to the Restriction Map Setup dialog 3 Click on OK in the Restriction Map Setup dialog The restriction enzymes and their binding sites will be shown in the Graphics Pane and Sequence Pane The specific cut site of each enzyme will be listed under Restriction Methylation Map in the Text Pane Analyses Cloning Gel An
11. GAACCT TACTAGATIG TICTIGTAC C a ie CCCACTTGGA F AGA GET AGGT ACTTAATCAC TCTATTGTCT CA TG ATCTCTCTTA CA p eT TCTTTCCTTA GTTAAAACTA TCATCCAATC ACAAGATGCG GGCTGGAAAG ACTTGCTCCC GAAGGATAAT CTTCTGCTT b AGAAAGGAAT CAATTTTGAT AGTAGGTTAG TGTTCTACGC CCGACCTTTC TGAACGAGGG CTTCCTATTA GAAGACGAA o AE np CCTCATATGd 310 bp 310bp CAGGG CTCATGCCCC TTCTTCCTTC GAACTGCCCG ATGAGGAAGT CCTTAGCCTA TCAAAGAATT CGGGACCAT Lel vand Pee Tet hel Dad jen TBN Dan mel meel ied ca a mam pDmma mmm RE Tat Bla al NN a ane het smd Tel on a at ales oel The ie oel het hol in a CF Sees ar A aaia a a a i a E pn penn ag Figure6 Selecting a DNA sequence by 1 dragging in Graphics Pane or 2 dragging in Sequence Pane To copy a molecule sequence 1 Select it as described above 2 To copy it to the Windows clipboard use the CTRL C keyboard command or To copy the sequence as a text file go to the Edit menu and select Copy to File You will be prompted to select a format and enter a name for the file To delete a molecule sequence 1 Select it as described above 2 Click on the DELETE key on your keyboard To paste a molecule sequence 1 With the sequence in text format on the Windows clipboard click on the point in the Sequence Pane where you want to add the insert 2 Click on CTRL V on your keyboard 3 The Insert Sequence dialog will open displaying the sequence to be inserted 4 Click on OK to complete the insertion Designing PCR Primer
12. GGCGGAAA AAGGGCTCCC ACCCCETCTT GGCATATATT CACGTCATCA GCGGCACTTG CAAGAAAAAG CGTTGCCCAA n NAAR 301 GTGCCGTGTG TGGTTCCCGC GGGCC I GGCC TCTTTACGGG TTATGGCCCT TGCGTGCCTT GAATTACTTC CACCTGGCTG Y E Al b Ready Lbp xi Rest site shown in Sequence Pane Figure 13 Creating a Restriction Map Aligning Molecules Vector NTI Advance 11 5 can align the sequences of two or more DNA RNA molecules The tool for doing this is called AlignX This tool can be launched from either the Molecule Viewer or Vector NTI Explorer To align sequences using Vector NTI Explorer 1 In the Explorer select the molecules that you want to align using SHIFT CLICK or CTRL CLICK key commands Figure 14 2 Go the Align menu and select AlignX Align Selected Molecules The AlignX Window will open with the molecules you selected listed in the upper left Text Pane 3 In the AlignX Window use SHIFT CLICK or CTRL CLICK key commands to select two or more molecules in the Text Pane list to align 4 To begin the alignment click on the Align button on the toolbar The alignment may take several minutes depending on the length and number of the molecules selected 5i When the alignment is complete the results are displayed in the AlignX Window as shown in Figure 14 The AlignX Window has panes showing different similarity graphs and the points at which the sequences align m Expl
13. Invitrogen by technologies Vector NTI Advance 11 5 Quick Start Guide Catalog no 12605050 12605099 12605103 Part no 12605 022 Revision date 10 October 2010 MANO0000419 2010 Life Technologies Corporation All rights reserved For Research Use Only Not intended for any animal or human therapeutic or diagnostic use The software described in this document is furnished under a license agreement Life Technologies Corporation and its licensors retain all ownership rights to the software programs and related documentation Use of the software and related documentation is governed by the license agreement accompanying the software and applicable copyright law The trademarks mentioned herein are the property of Life Technologies Corporation or their respective owners GENEART is a registered trademark of GENEART AG Windows and Windows Vista are registered trademarks of Microsoft Corporation Macintosh Mac and Mac OS are registered trademarks of Apple Inc Life Technologies reserves the right to make and have made changes without notice both to this publication and to the product it describes Information concerning products not manufactured or distributed by Life Technologies is provided without warranty or representation of any kind and neither Life Technologies nor its affiliates will be liable for any damages Limited Use Label License No 358 Research Use Only The purchase of this product conveys to the purchaser t
14. Reading Frames ORFs ORFs in it based on start and stop codons within the molecule Vector NTI Advance 11 5 can analyze a DNA RNA molecule and identify the open reading frames 9 Analyses Cloning Gel Analysis List With a DNA or RNA molecule open in the Molecule Viewer 1 2 Go to the Analyses menu and select ORF Figure 12 In the dialog box select the parameters for identifying and marking ORFs in the molecule When you click on OK the sequences identified as ORFs will be marked with directional arrows in the Graphics Pane and Sequence Pane and the ORFs will be listed in the Text Pane To identify an ORF in the different panes Click on a directional ORF arrow in the Graphics Pane to highlight its sequence in the Sequence Pane or Open a folder under Open Reading Frames in the Text Pane right click on the ORF name and select Find ORF to highlight it in the Graphics and Sequence Panes To save an ORF to the feature map of the molecule right click on the ORF arrow in the Graphics Pane or the ORF folder in the Text Pane and select Add ORF to FMap Thermodynamic Properties Oligo Duplexes y Wester NTI pci E m E lee GelAnaysis Lu Windy An Ambe Toos peip 85x ini e T LD Wn n 8 e ALFA ie Ea e SM GTE RO EEE A o em vi a Un 52 Eo 6 i4 qam id It ee en en A EN Find Motifs
15. Tod S60 G7 50 7 E a bmp arte hen Fmi Tand Siirat Primi Apa Liar Lergib 20 Tm 43 1 C DE 40 0 iby Pigeon of Arien Faua Lee B T TTGTAGTCTTGCAACATGO pc ari i di fom Bal bp Tar Sd be Mac BE bp Mar EAD bp Tailor y 300 9 2 e s Hammon Maris of Dupa Pimi EH 138 4 kcal mal d 355 Leland ek DD old irm Condi Andheri Primer icillin i m 0 Probe Ae 2 AU iS SAAR itm cs F ipii Sankey VOUS f pleni 3v TORD we Ta Cl tht Length Lemah 20 Ten 46 6 7 GC 55 0 weil b ET en mae a Thai te Ali 5r op pup o ESSE Ata ee enews eH 1545 B ecard i 2812 7 altered cs 532 Oral il mute aiiai x ian Vx r Tro Calter 3 8 GE Dhiman S 4 V ga fired Frame LATGTACTCT TATOCAATAC HW Gerne TTAGATCAGA ATACGTLATG anton Aeh to T emmae a arae Pima al Ardon Pes Lead eres Deda UK 0 at ipee 4 Raad arbe ti bp GAU bp Ll Figure 7 Designing PCR primers within a selected region To save the PCR analysis results as a separate object in the database 1 Right click on the PCR Analysis folder in the Text Pane 2 Select Save as Analysis Result The saved results will be listed under the Analysis Results object type in the Vector NTI Explorer Figure 8 Ue Exploring Local Vector NTI Database AL Analyses Edit View Assemble Tools Analyses Align Database Help TALAL 9 s 6 c B Analysis Results All database Analysis Results a Analysis Results MAIN Name Z PCR Analysi
16. a A i Heal aah j a EE m F fy CMU forward primer Amp Fa a ul Cuv Tarwras d piima ue M primer x A 40 TF pima m TP promoter En 25 Ti promet ie NEW FEATURE Jj i i E pli orgia r M ar Ms meenr ona tate bg A DEMO vector or CES M ped Ke gah wl 1 X VED Ian Ik BCE AU E zaad bp M Ni f A animes i M Ecc RE wala i i A melt AM J MMGH reverse primer i Nena Pir gery Jd Emi a e DEN p K 1 j Fragment Syp h PW anain i Let 57 fewer DUM Pd CT fares ij Us BW early promater VA Mest ita D mA Open Mec den Akt Fr agent n Y Tenus Lat Hi rere edo Higor To emet a fragment cick on the hate of De restriction erpen af the Tat Terman hold down he Shift bey ta Figure 10 Clone 2 fragments with Clone2Seq 11 In Silico Gene Synthesis with ReGENerator ReGENerator offers the fastest way to construct a DNA sequence based on the final protein molecule that you want to express You can introduce amino acid mutations and specify the flanking sequences you need for expression purification or detection When you are finished you can send the DNA sequence directly to the GENEART secure website www geneart com vectornti for rapid gene synthesis With a protein molecule loaded in the Molecule Viewer select Cloning gt ReGENerator or click on the S icon on the main toolbar to open ReGENerator Using the tool you can e Mutate the protein molecule by selecting amino acids in the Protein to
17. alysis List Thermodynamic Properties Oligo Duplexes Find Motifs T Find ORE Silent Mutations Translation Q Restriction Sites b gt Restriction Analyses Back Translation BioAnnotator GenomBench Tools Run Analysis Monitor Web Analyses wT r Restriction Fragments Restriction Fragment Length F Find Common Non Cutting Enz Restriction Report Restriction Map Setup Use Enzymes Permitted Terminus Types Ignore RENs cutting outside region v amp v 3 4 Blunt Add eK Remove Cancel gt gt Remove All Sart Sites in the Text Pane by Enzyme Name Number of Recognition Sites lanore RENs having less than sites more than sites Ignore RENs cutting inside region From To From To Choose Database Enzymes 2 xl huckieo Man San ir Look in Enzymes MAIN x Recognition Sting Terminus Type amp AarI cacctgc 5 Overhang mi Sat I gacate 3 Overhang 5 Acc6SI gatacc 5 Overhang mi Acct gtmkac 5 Overhang Acen cagctc 5 Overhang i Acil cege 5 Overhang lach aacgtt 5 Overhang fi Acur ctgaag 3 Overhang afer agcgct Blunt HJA cttaag 5 Overhang el AFIIII acrygt 5 Overhang m Age accggt 5 Overhang xj Select Al T Cancel 14 j Sif Vector NTI pTarget DEST meam si File Edit View Primer Design Analyses Cloning Gel Analysis List Window Align Assemble Tools He
18. emens nnmma ms nm i eemam 4l m sfa m oooo oo ce dt CAA T T N T C GAT GAT G G T T GA G AT tow 1600 ONETIKAN R 0 CA amp T TN T C GA T GAT G GT T GA G AT e 1 2 3 4 5 6 7 8 3 10 1 12 13 14 15 16 17 18 13 20 21 22 23 a em 4 uu gt d a a a a a Fragment Viewer History View 14 item s in Assembly 1 B mm on Name Length Original length 5 Trimmed bases i embly 3675 PGAR Crs diei a IUE TUM 737 759 i pm At p p Andres hee mE Rude ce zm ZqONE13SKANR 723 819 IGONEISKANF 567 784 S ONELOKANF 688 758 IGONEI1KANR 698 755 S ONELEKANF 599 767 IGONESKANR 719 758 eta zudem n eit iae Ma penema is ba GONEGKANR 714 770 Last PU aas Pai L5 D Pen cesa DID ceni 4 Verba vin bp emmen hmmm vmma nap tbid e ra 2m edle Zes ONESKANR 704 755 EEn GONEZKANR 705 747 3 Contig 2 764 EN u ONESKANR 762 764 a GONE4KANR 719 756 Mate ad hagen 13 ea DE Lil Ty Du p AME je Iu Gn cA fai s cp TEN j PON Conil ee OME LO LT EL Cooper tar CEL Ih jum s Fil Dai dnf CP 22580 oe OMA A74 TRAP gm ir p ERR Ap HEE CAE DATI a HET iur Fumi Pi a OA Cm ie jan rf SSE Ta fr 4 am A ASAP A Nall i NAT 1 hg it ne 1 f hy us JA n j V i i if M ry A Ws d WT fy nu i Plu T l T t a nc hkaaudcttctAdgtASM 3 T d asa imi 5 T4o a gt ak da
19. he limited non transferable right to use the purchased amount of the product only to perform internal research for the sole benefit of the purchaser No right to resell this product or any of its components is conveyed expressly by implication or by estoppel This product is for internal research purposes only and is not for use in commercial applications of any kind including without limitation quality control and commercial services such as reporting the results of purchaser s activities for a fee or other form of consideration For information on obtaining additional rights please contact outlicensing dlifetech com or Out Licensing Life Technologies 9791 Van Allen Way Carlsbad California 92008 Table of Contents MEO CLUE doe EN 4 Opening Vector NT Advance T 1 5 envel meiden neee deed sea ces 5 Moro B Database eet c EOM 6 Database Backup RestoF6e nannieb 7 MiOUC CUE ril 8 Selecting and Editing Molecule Sequences cccccecceeceeeeeeeeeeeeeaeeeaeeeeeeseeesenesauesaeeseeeseeeseesseeeaaes 9 Designing PCR Primers from a Sequence un nn nnnn enn enne nne ennn E ners a a ERE 10 Cloning Two Fragments with Clone2Seq nnunar snars neren senen enen enen ane vene nnn nana sana nan nn 11 In Silico Gene Synthesis with ReGENerator eoi desi rye ober kV Edo EYFMERU ci sun p ERE Eck Ec S Een E Ru pi ui 12 Identifying Open Reading Frames ORFS nennen nnee nee neee eerenenenenevereverenenvnenenenenenenen 13 Creat
20. ing a Restriction MAD aester aeebeekedenkidohedente au iubeat oon uo EEEREN 14 Abn MOVE CULES erie C E E ann bandana 15 Contig ASS CITI UY eer 16 TeenhicakSUPPOrbs edentate ennn eenden dee eehter eene se ennen enten 18 Introduction This Quick Start Guide is designed to get you started using Vector NTI Advance 11 5 It provides brief descriptions of the Vector NTI Advance 11 5 graphical user interface including Vector NTI Explorer and the Molecule Viewer and step by step instructions for using the most common features and functions of the software The topics covered include locating the desired tools displaying molecules designing PCR primers cloning two fragments gene synthesis aligning molecules performing a restriction analysis and assembling contigs This guide assumes that you have a working knowledge of basic Microsoft Windows and Mac OS features and functions how to open and save files how to use your mouse and so on and that Vector NTI Advance 11 5 is installed on your computer Opening Vector NTI Advance 11 5 The QuickStart Page is a single page that consolidates most commonly used modules tools and utilities that Vector NTI provides To launch the QuickStart Page select Start gt All Programs gt Invitrogen gt Vector NTI Advance 11 gt Quick Start r Vector NTI Advance 11 5 B QuickStart Page El E Launch Molecule Viewer Launch Local Database Assemble Chromatoaram Data a ta
21. ln ccr adgd ci os u i amp ak mns e 1 x x Me p w sar eed i E kn z v ATA GmugimrTacenrTraTcc ci sd rc eeecalaacacaasdad ddacatlatobl taabiageccraidca i a zi i Figure 15 Assembling a Contig 17 Technical Support Free technical support for Vector NTI Advance 11 5 is available exclusively through the web For more information check out the Software Support section at www invitrogen com VectorNTI To obtain personalized technical support by telephone or email you must have an annual support contract Users may purchase an Advanced Support Contract by contacting Invitrogen at bioinfosales lifetech com For paid support use the following contacts North America Phone 800 955 6288 x 67990 E mail bioinfosupport lifetech com Europe Middle East Africa Asian Pacific Phone 44 781 696 2707 Email bioinfosupport lifetech com 18 invitrogen vy Life technologies Corporate Headquarters 5791 Van Allen Way Carlsbad CA 92008 T 1 760 603 7200 F 1 760 602 6500 E tech_support invitrogen com For country specific contact information visit our web site at www invitrogen com
22. lp er oC uu O El s i C General Description EcoRI 5552 BamHI 4 O Standard Fields BamHI 5345 smal 11 r i CN aerem MANC S a Annotations EcoRI 5095 BamHI 25 S Restriction Methylation Map x C C EcoRI s E C wr GEATCC S L CJ BamHI 4 sites ecTack uc nsi ATEGAT Ll clat 0 sites TAGCIA ao 7 ci GhATTC EcoRI 7 sites crraak zd us cfe AAGCTT Fel CJ HindIII 1 site TTCGA mmm EcoRI 1251 Ncol 4794 w Pal 383 BamHI 4643 a Smal 817 PstI 888 a es EESTE Target DEST we om NcoI 1 site eee Smal 4229 P g pue Pstl 1260 fs 5583 bp N LL PstI 7 sites Er Ps 3998 EcoRI 1313 fla WL Smal 3 sites Mis EE EcoRI 3832 _ EcoRI 1624 o amp Invitrogen products related to pTarget DEST Pal 383 y Ps 1633 amp C Author attt Rest site shown in _ Original Author Bind 396 G r ap h ics P ane AZ Comments 4 illl b si Smal a PALAS BamHI BamHI EcoRI LAAD hhh ahh SISSIES VLAD 1 p dd GGGTACCGAG NNGGATCH CTAGTAACGG CCGCCAGTGT GCTGGAATTC GGCTTGGCTC CGGTGCCCGT TCCTAGGGG CCCATGGCTC NAGCCTAGGT GATCATTGCC GGCGGTCACA CGACCTTAAG CCGAACCGAG GCCACGGGCA i01 GCCCACAGTC CCCGAGAAGT TGGGGEGAGG GGTCGGCAAT TGAACCGGTG CCTAGAGAAG GTGGCGCGGG GTAAACTGGG CGGGTGTCAG GGGCTCTTCA ACCCCECTCC CCAGCCGTTA ACTTGGCCAC GGATCTCTTC CACCGCGCCC CATTTGACCC 201 CTCCGCCTTT TTCCCGAGGG TGGGGPAGAA CCGTATATAA GTGCAGTAGT CGCCGTGAAC GTTCTTTTTC GCAACGGGTT GA
23. mble Tools Help ieee tools a SY ER aa SE IK R R S e CARES 3 2 Tutorial 1 C General Description ApaL1 178 O C Standard Fields P BLA Fi PAP al gl Comments o Annotations Text Pane J Component Fragments info about E Cy 1 FRAGMENT of parent position from 415 to 396 features original length 2668 r molecule position from 1828 to 396 z restriction ER Left Terminus Tutorial 1 sites etc Smal ste 1 mm Graphics CyRight Terminus 4 p EcoRI site 1 Pan e Cy 2 FRAGMENT of parent position from 4360 to 1425 fill of 4 X original length 1431 we molecule position from 397 to 1827 Apali 2524 E Cy Left Terminus EcoRI site 1 5 Cy Right Terminus Avalsite 1 Completely filled in P LAC Feature Map C Restriction Methylation Map ApaL 3780 _ EcoRI 397 Cia1 423 Sana 428 P2P APr BamHi 774 Bambi 1831 Y ae ee PstI 1853 Hindili 1851 CGCTITCGCT GGAGCGCGAC GATGATCGGC CTGICGCITG CGGTATTCGG AATCTTGCAC GCCCTCGCTC AAGCCTTCGT CACTGGICCC GCCAC 4 GCGAAAGCGA CCTCGCGCTG CTACTAGCCG GACAGCGAAC GCCATAAGCC TTAGAACGIG CGGGAGCGAG TICGGAAGCA GIGACCAGGG CGGIG Sequence p Pane Ji GTTTCGGCGA GAAGCAGGCC ATTATCGCCG GCATGGCGGC CGACGCGCTG GGCTACGICT TGCIGGCGIT CGCGACGCGA GGCTGGATGG CCTTC CAAAGCCGCT CIICGICCGG TAATAGCGGC CGTACCGCCG GCIGCGCGAC CCGATGCAGA ACGACCGCAA GCGCTGCGCT CCGACCTACC GGAAG Ready
24. olecules Display analyze molecules Import analysis results ee p Export molecules o d LI P Import BLAST results Y Rd Launch tools LX ES E ANNAE VINE Ner TIR ME AEN Protein Molecules All database Protein Molecules Ii Praten Maglie PUSH Kare length Tonga Typa Jusihar Original Anhar Made A List of r AA H database SWISSPROT VANE 18 28 records 3 El ngos m ue Database Nn a object type BLAST Repu T Annhysis Prstira n oxi ee T Figure 2 Vector NTI Explorer Local Database window Database objects in Vector NTI Advance are categorized by type DNA molecules protein molecules and so on Some molecules are installed with the software When you first open the software DNA RNA Molecules is the selected object type Click on the tab in the lower left corner of the Vector NTI Explorer to select from the other available database objects To open an object from the local database double click on the object name in the right hand pane of the Explorer Depending on the object type information about that object may be displayed in a dialog box or the object may be loaded into a viewer For example DNA RNA and protein molecules are displayed in the Molecule Viewer When you install Vector NTI Advance the default local database is created in a Note folder called VNTI Database in the root directory of your computer e g
25. oring Lacal Vector NTI Database i ET Align Database Help C ich eng Edit View pcd Tools Analyses Align asd Help i D Al el IB t dis ci DE ec X e Alignx Open New Alignment Window DNA RNA Molecules All database DNA RNA Molecules Align Blocks Align Selected Molecules im TE DNA RNA Molecules MAIN Name Length Fom A 7 BP Invitrogen vectors amp pRSVNeo 5736 Circular Alignx Blacks Open New Alignment Window fall Cluster unclusterized amp pRSvneo 5736 Circular Web Alignment Tools gt E Sequencing Projects amp pscio1 9263 Circular v fall Bacterial Genomes amp pSCREEN iT lacZ DEST verA 8702 Circular iH Duplicates amp pSCREEN iT lacz GW CDK2 verA 7947 Circular amp pSELECT 1 5680 Circular 3176 Linear 7 i E oO 2999 Circular i mr 3005 Circular idi 2417 Circular MERE UNE mec aaa a 2419 Circular Hs ut Pg BEL b a A o 2a DNA RNA Mol 2462 Circular en AG es E nu o Xi 2464 Circular HOEN HN Be 5 EIC se 5003 Circular sonnet 8 parie zona 4900 Circular ern n D paras nns 5294 Circular KM Ja Epson 5503 Circular e fo paer 2419 el u E perte iz 6 DNA RNA molecules selected Vv ja E perri tzaa ui Untitled AlignX TUNES x Project Edit View Align Analyses Assemble Tools Window Help F Ansy emma n pribora DUP idem L EA SND Similarity Ei psP64 2999 1 amp amp psP65 3005 amp psp7o 2417 4 8 psP71 2419 E psP72 2462 8 pSsP7
26. rm contig assembly select the fragments in the ContigExpress Project Explorer kd Click the Assemble Selected Fragments icon S on the main toolbar Fragments will be analyzed and assembled into one or more contigs which will be listed in the Project Viewer along with the fragments in each contig Double click on a contig in the list It will be displayed in the Contig Viewer The Sequence Pane at the bottom shows the sequence of the assembly The Graphics Pane on the right shows the orientations of the fragments in the assembly The Text Pane on the left lists the fragments in the assembly If you wish to edit the contig enable the Enhanced Edit Mode by clicking the icon Use Enhanced Edit Mode far left on the toolbar in the Contig window before making any reasonable changes There are three trimming options in ContigExpress Fragments can be trimmed for ambiguities Phred quality scores and vector contamination Refer to the Vector NTI Advance 11 5 User s Manual for details Continued on the following page Contig Assembly continued Open Project Edit View Assemble Align Ctrl O Close Project Ctrl S Save Save Bs ants ts T zamelen QU abi El TwOLIR abs Revert to Saved Project From GenBank file Export Item From FASTA file Discard All Chromatograms TOE Dissolve Contig From SCF file My Meret Maree Convert Contig to Fragment From
27. s from a Sequence Vector NTI Advance 11 5 can analyze a selected sequence and design PCR primers for it based on parameters such as desired melting temperature Tm GC content and amplicon length With a DNA or RNA molecule open in the Molecule Viewer 1 Select the part of the sequence for which you want to design primers as described on the previous page 2 Go to the Primer Design menu J Select Find PCR Primers Inside Selection to find primers within the sequence Figure 7 or Select Amplify Selection to find primers in the regions before and after the sequence other amplification selections are available see the Vector NTI Advance 11 5 User s Manual for more information 4 In the dialog box select the desired primer design parameters Note that most of these parameters have default values based on typical PCR primers 5 Click on OK The results will appear under PCR Analysis in the Text Pane Primer Design Analyses Cloning Gel Analysis Find PCR Primers Inside Selection ie vector WE puert STOP C vera Tee Amplify Selecti R He Ed yew Primer Design Anases Coney Geldnalyels Wet Window dgn Assemble Toos mpil election ii PCR analysis tre ex i ea i EA og pum ri Eh wn results un BI ae TTE EURO c ns BN mE 0 E prea Analysis P wv JY at Prod E al bert 517 ralin 171 R amp V pri hann ee CIENT Cronica region nf Ehe maande Ienm 21 tn 5 His IEEE i psy mare n Fis iei Falke j Tre MAC
28. s of M13 iai e le a es 1 analysis result HYP a Figure 8 PCR analysis results listed in Vector NTI Explorer Cloning Two Fragments with Clone2Seq You can use Clone2Seq to easily clone two fragments in Vector NTI Advance 11 5 1 In the Molecule Viewer go to Cloning gt Clone2Seq or click on the Clone2Seq button on the main toolbar 2 To load a molecule e g an insert or vector click on Open Molecule in one of the window panes 3 With the desired restriction enzyme sites displayed click on the button to select select the first restriction site by clicking on it then hold down the SHIFT key and click on the second site TAI 203 Asal 298 AsnI 298 Asal 239 AsIGso jj Shift click to select the second restriction site Protein Bind 1 35 Signal 1 10 Signal 1 Hiinal 9 QAI pUCi9L 2659 bp Molecule 1 Open Molecule Add Fragment 5 Terminus 3 Terminus Site PvulI 1 at 192 bp Site Pvul 2 at 2527 bp Edit Fragment Clear Fragment Figure9 Selecting restriction sites in the Clone2Seq viewer 4 Add the selected fragment by clicking on Add Fragment 5 Repeat these steps with the second molecule Make sure the left terminus of the first fragment is compatible with the right terminus of the second fragment and vice versa 6 When you have selected both fragments click on Clone mmm bla picmadnr Oll genten L fa j ETT eP peli ige
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