LOCUS       Exported               12543 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  Plant transient expression vector pTK251, complete sequence.
ACCESSION   JX971625
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 12543)
  AUTHORS   Kagale S, Uzuhashi S, Wigness M, Bender T, Yang W, Borhan MH, 
            Rozwadowski K.
  TITLE     TMV-Gate vectors: Gateway compatible tobacco mosaic virus based 
            expression vectors for functional analysis of proteins
  JOURNAL   Sci Rep 2, 874 (2012)
  PUBMED    23166857
REFERENCE   2  (bases 1 to 12543)
  AUTHORS   Rozwadowski K, Kagale S.
  TITLE     Direct Submission
  JOURNAL   Submitted (15-OCT-2012) Molecular Genetics, Agriculture and 
            Agri-Food Canada, 107 Science Place, Saskatoon, Saskatchewan S7N 
            0X2, Canada
REFERENCE   3  (bases 1 to 12543)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     ##Assembly-Data-START##
            Sequencing Technology :: Sanger dideoxy sequencing 
            ##Assembly-Data-END##
FEATURES             Location/Qualifiers
     source          1..12543
                     /organism="Plant transient expression vector pTK251"
                     /mol_type="other DNA"
                     /db_xref="taxon:1261656"
     misc_feature    5..29
                     /label=LB T-DNA repeat
                     /note="left border repeat from nopaline C58 T-DNA"
     misc_feature    complement(82..12469)
                     /label=left border
                     /note="left border"
     primer_bind     110..126
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     regulatory      146..899
                     /regulatory_class="promoter"
                     /note="CaMV 35S"
     promoter        854..899
                     /label=minimal CaMV 35S promoter
                     /note="minimal 35S promoter from cauliflower mosaic virus"
     misc_feature    911..966
                     /label=TMV Omega
                     /note="translational enhancer from the tobacco mosaic virus
                     5-leader sequence (Gallie et al., 1988)"
     CDS             968..4318
                     /codon_start=1
                     /gene="Rep"
                     /product="replicase"
                     /label=Rep
                     /protein_id="AFX83519.1"
                     /translation="MAYTQTATTSALLDTVRGNNSLVNDLAKRRLYDTAVEEFNARDRR
                     PKVNFSKVISEEQTLIATRAYPEFQITFYNTQNAVHSLAGGLRSLELEYLMMQIPYGSL
                     TYDIGGNFASHLFKGRAYVHCCMPNLDVRDIMRHEGQKDSIELYLSRLERGGKTVPNFQ
                     KEAFDRYAEIPEDAVCHNTFQTCEHQPMQQSGRVYAIALHSIYDIPADEFGAALLRKNV
                     HTCYAAFHFSENLLLEDSCVNLDEINACFSRDGDKLTFSFASESTLNYCHSYSNILKYV
                     CKTYFPASNREVYMKEFLVTRVNTWFCKFSRIDTFLLYKGVAHKSVDSEQFYTAMEDAW
                     HYKKTLAMCNSERILLEDSSSVNYWFPKMRDMVIVPLFDISLETSKRTRKEVLVSKDFV
                     FTVLNHIRTYQAKALTYANVLSFVESIRSRVIINGVTARSEWDVDKSLLQSLSMTFYLH
                     TKLAVLKDDLLISKFSLGSKTVCQHVWDEISLAFGNAFPSVKERLLNRKLIRVAGDALE
                     IRVPDLYVTFHDRLVTEYKASVDMPALDIRKKMEETEVMYNALSELSVLRESDKFDVDV
                     FSQMCQSLEVDPMTAAKVIVAVMSNESGLTLTFERPTEANVALALQDQEKASEGALVVT
                     SREVEEPSMKGSMARGELQLAGLAGDHPESSYSKNEEIESLEQFHMATADSLIRKQMSS
                     IVYTGPIKVQQMKNFIDSLVASLSAAVSNLVKILKDTAAIDLETRQKFGVLDVASRKWL
                     IKPTAKSHAWGVVETHARKYHVALLEYDEQGVVTCDDWRRVAVSSESVVYSDMAKLRTL
                     RRLLRNGEPHVSSAKVVLVDGVPGCGKTKEILSRVNFDEDLILVPGKQAAEMIRRRANS
                     SGIIVATKDNVKTVDSFMMNFGKSTRCQFKRLFIDEGLMLHTGCVNFLVAMSLCEIAYV
                     YGDTQQIPYINRVSGFPYPAHFAKLEVDEVETRRTTLRCPADVTHYLNRRYEGFVMSTS
                     SVKKSVSQEMVGGAAVINPISKPLHGKILTFTQSDKEALLSRGYSDVHTVHEVQGETYS
                     DVSLVRLTPTPVSIIAGDSPHVLVALSRHTCSLKYYTVVMDPLVSIIRDLEKLSSYLLD
                     MYKVDAGTQ"
     gene            968..4318
                     /gene="Rep"
                     /label=Rep
     CDS             4394..5812
                     /codon_start=1
                     /gene="RNA pol"
                     /product="RNA polymerase"
                     /label=RNA pol
                     /protein_id="AFX83520.1"
                     /translation="MQFYYDKCLPGNSTMMNNFDAVTMRLTDISLNVKDCILDMSKSVA
                     APKDQIKPLIPMVRTAAEMPRQTGLLENLVAMIKRNFNAPELSGIIDIENTASLVVDKF
                     FDSYLLKEKRKPNKNVSLFSRESLNRWLEKQEQVTIGQLADFDFVDLPAVDQYRHMIKA
                     QPKQKLDTSIQTEYPALQTIVYHQKRSMQYRPLLVSLPGNYWTVLIRADFVFTRKTPAQ
                     IEDFFGDLDSHVPMDVLELDISKYDKSQNEFHCAVEYEIWRRLGFEDFLGEVWKQGHRK
                     TTLKDYTAGIKTCIWYQRKSGDVTTFIGNTVIIAACLASMLPMEKIIKGAFCGDDSLLY
                     FPKGCEFPDVQHSANLMWNFEAKLFKKQYGYFCGRYVIHHDRGCIVYYDPLKLISKLGA
                     KHIKDWEHLEEFRRSLCDVAVSLNNCAYYTQLDDAVWEVHKTAPPGSFVYKSLVKYLSD
                     KVLFRSLFIDGSSC"
     gene            4394..5812
                     /gene="RNA pol"
                     /label=RNA pol
     CDS             5796..6602
                     /codon_start=1
                     /gene="MP"
                     /product="movement protein"
                     /label=MP
                     /protein_id="AFX83521.1"
                     /translation="MALVVKGKVNINEFIDLTKMEKILPSMFTPVKSVMCSKVDKIMVH
                     ENESLSGVNLLKGVKLIDSGYVCLAGLVVTGEWNLPDNCRGGVSVCLVDKRMERADEAI
                     LGSYYTAAAKKRFQFKVVPNYAITTQDAMKNVWQVLVNIRNVKMSAGFCPLSLEFVSVC
                     IVYRNNIKLGLREKITNVRDGGPMELTEEVVDEFMEDVPMSIRLAKFRSRTGKKSDVRK
                     GKNSSSDRSVPNKNYRNVKDFGGMSFKKNNLIDDDSEATVAESDSF"
     gene            5796..6602
                     /gene="MP"
                     /label=MP
     misc_feature    6683..6765
                     /label=StrepTag_N-terminal fusion
                     /note="StrepTag_N-terminal fusion"
     CDS             6683..6706
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             6743..6766
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     misc_feature    6800..6817
                     /label=6xHis
                     /note="6xHis"
     CDS             6800..6817
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             6821..6853
                     /codon_start=1
                     /product="leader peptide from bacteriophage T7 gene 10"
                     /label=T7 tag (gene 10 leader)
                     /note="promotes efficient translation in E. coli"
                     /translation="MASMTGGQQMG"
     misc_feature    6857..6880
                     /label=Xpress epitope
                     /note="Xpress epitope"
     CDS             6857..6880
                     /codon_start=1
                     /product="Xpress(TM) epitope tag, including an enterokinase
                     recognition and cleavage site"
                     /label=Xpress(TM) tag
                     /translation="DLYDDDDK"
     misc_feature    6866..6880
                     /label=enterokinase protease recognition site
                     /note="enterokinase protease recognition site"
     misc_recomb     6890..7014
                     /label=attR1
                     /note="attR1"
     protein_bind    6890..7014
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        7039..7069
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     CDS             7123..7782
                     /codon_start=1
                     /gene="CmR"
                     /product="chloramphenicol resistance protein"
                     /label=CmR
                     /protein_id="AFX83522.1"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     gene            7123..7782
                     /gene="CmR"
                     /label=CmR
     CDS             8124..8429
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB"
                     /label=ccdB
                     /protein_id="AFX83523.1"
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     gene            8124..8429
                     /gene="ccdB"
                     /label=ccdB
     misc_recomb     8470..8594
                     /label=attR2
                     /note="attR2"
     protein_bind    complement(8470..8594)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     misc_feature    8839..8883
                     /note="TobRV/G12a_Ribozyme: aligns with Tobacco ringspot 
                     virus satellite RNA and Hammerhead ribozyme G12a"
     regulatory      8926..9136
                     /regulatory_class="terminator"
                     /note="CaMV 35S"
     misc_feature    complement(9166..9335)
                     /label=right border
                     /note="right border"
     misc_feature    9206..9230
                     /label=RB T-DNA repeat
                     /note="right border repeat from nopaline C58 T-DNA"
     CDS             complement(9382..10530)
                     /codon_start=1
                     /gene="RK2-TrfA"
                     /product="RK2-TrfA"
                     /label=RK2-TrfA
                     /protein_id="AFX83524.1"
                     /translation="MNRTFDRKAYRQELIDAGFSAEDAETIASRTVMRAPRETFQSVGS
                     MVQQATAKIERDSVQLAPPALPAPSAAVERSRRLEQEAAGLAKSMTIDTRGTMTTKKRK
                     TAGEDLAKQVSEAKQAALLKHTKQQIKEMQLSLFDIAPWPDTMRAMPNDTARSALFTTR
                     NKKIPREALQNKVIFHVNKDVKITYTGVELRADDDELVWQQVLEYAKRTPIGEPITFTF
                     YELCQDLGWSINGRYYTKAEECLSRLQATAMGFTSDRVGHLESVSLLHRFRVLDRGKKT
                     SRCQVLIDEEIVVLFAGDHYTKFIWEKYRKLSPTARRMFDYFSSHREPYPLKLETFRLM
                     CGSDSTRVKKWREQVGEACEELRGSGLVEHAWVNDDLVHCKR"
     gene            complement(9382..10530)
                     /gene="RK2-TrfA"
                     /label=RK2-TrfA
     CDS             complement(10829..11623)
                     /codon_start=1
                     /gene="KmR"
                     /allele="kanamycin resistance marker"
                     /product="kanamycin resistance protein"
                     /label=KmR
                     /protein_id="AFX83525.1"
                     /translation="MAKMRISPELKKLIEKYRCVKDTEGMSPAKVYKLVGENENLYLKM
                     TDSRYKGTTYDVEREKDMMLWLEGKLPVPKVLHFERHDGWSNLLMSEADGVLCSEEYED
                     EQSPEKIIELYAECIRLFHSIDISDCPYTNSLDSRLAELDYLLNNDLADVDCENWEEDT
                     PFKDPRELYDFLKTEKPEEELVFSHGDLGDSNIFVKDGKVSGFIDLGRSGRADKWYDIA
                     FCVRSIREDIGEEQYVELFFDLLGIKPDWEKIKYYILLDELF"
     gene            complement(10829..11623)
                     /gene="KmR"
                     /allele="kanamycin resistance marker"
                     /label=KmR
     rep_origin      complement(11835..12468)
                     /direction=LEFT
                     /label=RK2-oriV
                     /note="RK2-oriV"
     rep_origin      11835..12366
                     /label=oriV
                     /note="incP origin of replication"
     rep_origin      11843..12457
                     /label=oriV
                     /note="origin of replication for the bacterial F plasmid"