LOCUS       Exported               12872 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  Binary expression vector NES-YC3.6, complete sequence.
ACCESSION   JN903572
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 12872)
  AUTHORS   Krebs M, Held K, Binder A, Hashimoto K, Den Herder G, Parniske M, 
            Kudla J, Schumacher K.
  TITLE     FRET-based genetically encoded sensors allow high-resolution live 
            cell imaging of Ca(2) dynamics
  JOURNAL   Plant J. 69 (1), 181-192 (2012)
  PUBMED    21910770
REFERENCE   2  (bases 1 to 12872)
  AUTHORS   Krebs M, Held K, Binder A, Hashimoto K, Den Herder G, Parniske M, 
            Kudla J, Schumacher K.
  TITLE     Direct Submission
  JOURNAL   Submitted (24-OCT-2011) Department of Developmental Biology, Centre 
            for Organismal Studies Heidelberg, Im Neuenheimer Feld 230, 
            Heidelberg 69120, Germany
REFERENCE   3  (bases 1 to 12872)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..12872
                     /organism="Binary expression vector NES-YC3.6"
                     /mol_type="other DNA"
                     /db_xref="taxon:1154702"
     misc_feature    1..194
                     /label=right border
                     /note="right border"
     misc_feature    113..137
                     /label=RB T-DNA repeat
                     /note="right border repeat from nopaline C58 T-DNA"
     protein_bind    302..323
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        338..368
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    376..392
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     400..416
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     regulatory      444..1088
                     /regulatory_class="promoter"
                     /note="AtUBQ10"
     CDS             1114..3159
                     /codon_start=1
                     /product="NES-YC3.6"
                     /label=NES-YC3.6
                     /protein_id="AFC41195.1"
                     /translation="MLQNELALKLAGLDINKTGGVDSSAAATMVSKGEELFTGVVPILV
                     ELDGDVNGHRFSVSGEGEGDATYGKLTLKFICTTGKLPVPWPTLVTTLTWGVQCFSRYP
                     DHMKQHDFFKSAMPEGYVQERTIFFKDDGNYKTRAEVKFEGDTLVNRIELKGIDFKEDG
                     NILGHKLEYNYISHNVYITADKQKNGIKAHFKIRHNIEDGSVQLADHYQQNTPIGDGPV
                     LLPDNHYLSTQSALSKDPNEKRDHMVLLEFVTAARMHDQLTEEQIAEFKEAFSLFDKDG
                     DGTITTKELGTVMRSLGQNPTEAELQDMINEVDADGNGTIYFPEFLTMMARKMKDTDSE
                     EEIREAFRVFDKDGNGYISAAQLRHVMTNLGEKLTDEEVDEMIREADIDGDGQVNYEEF
                     VQMMTAKGGKRRWKKNFIAVSAANRFKKISSSGALELMDGGVQLADHYQQNTPIGDGPV
                     LLPDNHYLSYQSALSKDPNEKRDHMVLLEFVTAAGITLGMDELYKGGSGGMVSKGEELF
                     TGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKLICTTGKLPVPWPTLVTTLGYG
                     LQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDGNYKTRAEVKFEGDTLVNRIELK
                     GIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIKANFKIRHNIE"
     misc_feature    1114..1174
                     /label=NES
                     /note="NES"
     regulatory      1192..1201
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             2338..2415
                     /codon_start=1
                     /product="calmodulin-binding peptide"
                     /label=CBP
                     /note="derived from skeletal muscle myosin light chain 
                     kinase; binds calmodulin with nanomolar affinity in the 
                     presence of calcium"
                     /translation="KRRWKKNFIAVSAANRFKKISSSGAL"
     CDS             2638..3156
                     /codon_start=1
                     /product="N-terminal fragment of mVenus for use in 
                     bimolecular fluorescence complementation (BiFC) (Kodama and
                     Hu, 2010)"
                     /label=VN173
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KLICTTGKLPVPWPTLVTTLGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIK
                     ANFKIRHNIE"
     regulatory      3174..3820
                     /regulatory_class="terminator"
                     /note="rbcs"
     terminator      3175..3816
                     /label=E9 terminator
                     /note="terminator and polyadenylation signal from the pea 
                     rbcS-E9 gene"
     primer_bind     complement(3829..3845)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     regulatory      complement(4009..4700)
                     /regulatory_class="terminator"
                     /note="mas"
     terminator      4427..4679
                     /label=MAS terminator
                     /note="mannopine synthase terminator"
     CDS             complement(4875..5669)
                     /codon_start=1
                     /product="nptII"
                     /label=nptII
                     /protein_id="AFC41196.1"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     regulatory      complement(5699..6383)
                     /regulatory_class="promoter"
                     /note="mas"
     promoter        complement(5699..6079)
                     /label=MAS promoter
                     /note="mannopine synthase promoter (Velten et al., 1984)"
     misc_feature    6384..6712
                     /label=left border
                     /note="left border"
     misc_feature    6451..6475
                     /label=LB T-DNA repeat
                     /note="left border repeat from nopaline C58 T-DNA"
     CDS             6862..7794
                     /codon_start=1
                     /product="aadA"
                     /label=aadA
                     /protein_id="AFC41197.1"
                     /translation="MTCFFGVQSMPRASKQQARYAVGRCLMLWSSNDVTQQGSRPKTKL
                     NIMGEAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPHSDIDLLVTVTVR
                     LDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAKRELQFGEWQRND
                     ILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLFEALNETLTLWNS
                     PPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQPVILEARQAYLGQ
                     EEDRLASRADQLEEFVHYVKGEITKVVGK"
     CDS             7003..7794
                     /codon_start=1
                     /gene="aadA"
                     /product="aminoglycoside adenylyltransferase (Murphy, 
                     1985)"
                     /label=SmR
                     /note="confers resistance to spectinomycin and 
                     streptomycin"
                     /translation="MGEAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
                     SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
                     RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
                     EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
                     VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK"
     rep_origin      8038..8626
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     misc_feature    8812..8952
                     /label=bom
                     /note="basis of mobility region from pBR322"
     rep_origin      9296..9490
                     /label=pVS1 oriV
                     /note="origin of replication for the Pseudomonas plasmid 
                     pVS1 (Heeb et al., 2000)"
     CDS             complement(9556..10623)
                     /codon_start=1
                     /product="replication protein from the Pseudomonas plasmid 
                     pVS1 (Heeb et al., 2000)"
                     /label=pVS1 RepA
                     /translation="GRKPSGPVQIGAALGDDLVEKLKAAQAAQRQRIEAEARPGESWQA
                     AADRIRKESRQPPAAGAPSIRKPPKGDEQPDFFVPMLYDVGTRDSRSIMDVAVFRLSKR
                     DRRAGEVIRYELPDGHVEVSAGPAGMASVWDYDLVLMAVSHLTESMNRYREGKGDKPGR
                     VFRPHVADVLKFCRRADGGKQKDDLVETCIRLNTTHVAMQRTKKAKNGRLVTVSEGEAL
                     ISRYKIVKSETGRPEYIEIELADWMYREITEGKNPDVLTVHPDYFLIDPGIGRFLYRLA
                     RRAAGKAEARWLFKTIYERSGSAGEFKKFCFTVRKLIGSNDLPEYDLKEEAGQAGPILV
                     MRYRNLIEGEASAGS"
     CDS             complement(11057..11686)
                     /codon_start=1
                     /product="stability protein from the Pseudomonas plasmid 
                     pVS1 (Heeb et al., 2000)"
                     /label=pVS1 StaA
                     /translation="MKVIAVLNQKGGSGKTTIATHLARALQLAGADVLLVDSDPQGSAR
                     DWAAVREDQPLTVVGIDRPTIDRDVKAIGRRDFVVIDGAPQAADLAVSAIKAADFVLIP
                     VQPSPYDIWATADLVELVKQRIEVTDGRLQAAFVVSRAIKGTRIGGEVAEALAGYELPI
                     LESRITQRVSYPGTAAAGTTVLESEPEGDAAREVQALAAEIKSKLI"