LOCUS       40924_6966        8730 bp DNA     circular SYN 17-DEC-2018
DEFINITION  Cloning vector pBOB, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8730)
  AUTHORS   Wachsman G, Heidstra R, Scheres B.
  TITLE     Distinct Cell-Autonomous Functions of RETINOBLASTOMA-RELATED in 
            Arabidopsis Stem Cells Revealed by the Brother of Brainbow Clonal 
            Analysis System
  JOURNAL   Plant Cell 23 (7), 2581-2591 (2011)
  PUBMED    21742994
REFERENCE   2  (bases 1 to 8730)
  AUTHORS   Wachsman G, Heidstra R, Scheres B.
  TITLE     Direct Submission
  JOURNAL   Submitted (06-MAY-2011) Molecular Genetics, Utrecht University, 
            Padualaan 8, Utrecht 3584CH, The Netherlands
REFERENCE   3  (bases 1 to 8730)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 8730)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Plant 
            Cell"; date: "2011"; volume: "23"; issue: "7"; pages: "2581-2591"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (06-MAY-2011) Molecular Genetics, Utrecht University, Padualaan 8, 
            Utrecht 3584CH, The Netherlands"
COMMENT     SGRef: number: 3; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..8730
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             320..1132
                     /codon_start=1
                     /label=KanR
                     /note="aminoglycoside phosphotransferase"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANVVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     rep_origin      1306..1894
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     misc_feature    complement(1985..2009)
                     /label=RB T-DNA repeat
                     /note="right border repeat from nopaline C58 T-DNA"
     protein_bind    2248..2269
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     promoter        2284..2314
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2322..2338
                     /label=lac operator
                     /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     2346..2362
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        2383..2401
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        2485..2830
                     /label=CaMV 35S promoter
                     /note="strong constitutive promoter from cauliflower mosaic
                     virus"
     CDS             2907..2927
                     /codon_start=1
                     /label=SV40 NLS
                     /note="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /translation="PKKKRKV"
     CDS             2925..3641
                     /codon_start=1
                     /label=Venus
                     /note="yellow fluorescent protein (YFP) with fast and
                     efficient maturation (Nagai et al., 2002)"
                     /translation="VVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KLICTTGKLPVPWPTLVTTLGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIK
                     ANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     protein_bind    complement(4169..4202)
                     /label=lox2272
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     CDS             4275..4988
                     /codon_start=1
                     /label=CyPet
                     /note="cyan fluorescent protein optimized for FRET with
                     YPet (Nguyen and Daugherty, 2005)"
                     /translation="VSKGEELFGGIVPILVELEGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTWGVQCFSRYPDHMKQHDFFKSVMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYISHNVYITADKQKNGIKA
                     NFKARHNITDGSVQLADHYQQNTPIGDGPVILPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     terminator      5014..5266
                     /label=NOS terminator
                     /note="nopaline synthase terminator and poly(A) signal"
     protein_bind    complement(5296..5329)
                     /label=loxN
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AGGTATAC) (Shaw et al., 2021). loxN sites are 
                     compatible with each other, but incompatible with loxP or 
                     lox2272 sites."
     CDS             5399..6091
                     /codon_start=1
                     /label=TagRFP
                     /note="monomeric derivative of red fluorescent protein from
                     Entacmaea quadricolor (Merzlyak et al., 2007)"
                     /translation="GSELIKENMHMKLYMEGTVNNHHFKCTSEGEGKPYEGTQTMRIKV
                     VEGGPLPFAFDILATSFMYGSRTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLTA
                     TQDTSLQDGCLIYNVKIRGVNFPSNGPVMQKKTLGWEANTEMLYPADGGLEGRSDMALK
                     LVGGGHLICNFKTTYRSKKPAKNLKMPGVYYVDHRLERIKEADKETYVEQHEVAVARYC
                     DLPSKLGHK"
     promoter        complement(6384..6402)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(6412..6428)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        6646..6991
                     /label=CaMV 35S promoter
                     /note="strong constitutive promoter from cauliflower mosaic
                     virus"
     CDS             7024..7584
                     /codon_start=1
                     /label=DHFR
                     /note="mouse dihydrofolate reductase"
                     /translation="MVRPLNCIVAVSQNMGIGKNGDRPWPPLRNEFKYFQRMTTTSSVE
                     GKQNLVIMGRKTWFSIPEKNRPLKDRINIVLSRELKEPPRGAHFLAKSLDDALRLIEQP
                     ELASKVDMVWIVGGSSVYQEAMNQPGHLRLFVTRIMQEFESDTFFPEIDLGKYKLLPEY
                     PGVLSEVQEEKGIKYKFEVYEKKD"
     misc_feature    complement(8167..8189)
                     /label=LB T-DNA repeat
                     /note="left border repeat from nopaline C58 T-DNA
                     (truncated)"
     rep_origin      complement(join(8324..8730,1..29))
                     /direction=LEFT
                     /label=pSa ori
                     /note="origin of replication from bacterial plasmid pSa"
ORIGIN
        1 agcccgctcg acggcgggct gttggatggg gatcgcctga atcgccccat catccagcca
       61 gaaagtgagg gagccacggt tgatgagagc tttgttgtag gtggaccagt tggtgatttt
      121 gaacttttgc tttgccacgg aacggtctgc gttgtcggga agatgcgtga tctgatcctt
      181 caactcagca aaagttcgat ttattcaaca aagccacgtt gtgtctcaaa atctctgatg
      241 ttacattgca caagataaaa atatatcatc atgaacaata aaactgtctg cttacataaa
      301 cagtaataca aggggtgtta tgagccatat tcaacgggaa acgtcttgct caaggccgcg
      361 attaaattcc aacatggatg ctgatttata tgggtataaa tgggctcgcg ataatgtcgg
      421 gcaatcaggt gcgacaatct accgattgta tgggaagccc gatgcgccag agttgtttct
      481 gaaacatggc aaaggtagcg ttgccaatgt tgttacagat gagatggtca gactaaactg
      541 gctgacggaa tttatgcctc ttccgaccat caagcatttt atccgtactc ctgatgatgc
      601 atggttactc accactgcga tcccagggaa aacagcattc caggtattag aagaatatcc
      661 tgattcaggt gaaaatattg ttgatgcgct ggcagtgttc ctgcgccggt tgcattcgat
      721 tcctgtttgt aattgtcctt ttaacagcga tcgcgtattt cgtctcgctc aggcgcaatc
      781 acgaatgaat aacggtttgg ttgatgcgag tgattttgat gacgagcgta atggctggcc
      841 tgttgaacaa gtctggaaag aaatgcataa acttttgcca ttctcaccgg attcagtcgt
      901 cactcatggt gatttctcac ttgataacct tatttttgac gaggggaaat taataggttg
      961 tattgatgtt ggacgagtcg gaatcgcaga ccgataccag gatcttgcca tcctatggaa
     1021 ctgcctcggt gagttttctc cttcattaca gaaacggctt tttcaaaaat atggtattga
     1081 taatcctgat atgaataaat tgcagtttca tttgatgctc gatgagtttt tctaatcact
     1141 agaccaatgt tacacatata tactttagat tgatttaaaa cttcattttt aatttaaaag
     1201 gatctaggtg aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc
     1261 gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt
     1321 tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt
     1381 gccggatcaa gagctaccaa ctcttcttcc gaaggtaact ggcttcagca gagcgcagat
     1441 accaaatact gttcttctag tgtagccgta gttaggccac cacttcaaga actctgtagc
     1501 accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa
     1561 gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg
     1621 ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag
     1681 atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag
     1741 gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa
     1801 cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt
     1861 gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg
     1921 gttcctggcc ttttgctggc cttttgctca catgagatct caaacaaaca catacagcga
     1981 cttagtttac ccgccaatat atcctgtcaa ggccttcatg ttctttcctg cgttatcccc
     2041 tgattctgtg gataaccgta ttaccgcctt tgagtgagct gataccgctc gccgcagccg
     2101 aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa gagcgcccaa tacgcaaacc
     2161 gcctctcccc gcgcgttggc cgattcatta atgcagctgg cacgacaggt ttcccgactg
     2221 gaaagcgggc agtgagcgca acgcaattaa tgtgagttag ctcactcatt aggcacccca
     2281 ggctttacac tttatgcttc cggctcgtat gttgtgtgga attgtgagcg gataacaatt
     2341 tcacacagga aacagctatg accatgatta cgccaagcgc gcaattaacc ctcactaaag
     2401 ggaacaaaag ctggagctcc accgcggatc gtacccctac tccaaaaatg tcaaagatac
     2461 agtctcagaa gaccaaaggg ctattgagac ttttcaacaa agggtaattt cgggaaacct
     2521 cctcggattc cattgcccag ctatctgtca cttcatcgaa aggacagtag aaaaggaagg
     2581 tggctcctac aaatgccatc attgcgataa aggaaaggct atcattcaag atgcctctgc
     2641 cgacagtggt cccaaagatg gacccccacc cacgaggagc atcgtggaaa aagaagacgt
     2701 tccaaccacg tcttcaaagc aagtggattg atgtgacatc tccactgacg taagggatga
     2761 cgcacaatcc cactatcctt cgcaagaccc ttcctctata taaggaagtt catttcattt
     2821 ggagaggaca taacttcgta taggatactt tatacgaagt tattctagaa taacttcgta
     2881 tagtatacct tatacgaagt tatatgccaa agaagaagag aaaggtcgtg agcaagggcg
     2941 aggagctgtt caccggggtg gtgcccatcc tggtcgagct ggacggcgac gtaaacggcc
     3001 acaagttcag cgtgtccggc gagggcgagg gcgatgccac ctacggcaag ctgaccctga
     3061 agctgatctg caccaccggc aagctgcccg tgccctggcc caccctcgtg accaccctgg
     3121 gctacggcct gcagtgcttc gcccgctacc ccgaccacat gaagcagcac gacttcttca
     3181 agtccgccat gcccgaaggc tacgtccagg agcgcaccat cttcttcaag gacgacggca
     3241 actacaagac ccgcgccgag gtgaagttcg agggcgacac cctggtgaac cgcatcgagc
     3301 tgaagggcat cgacttcaag gaggacggca acatcctggg gcacaagctg gagtacaact
     3361 acaacagcca caacgtctat atcaccgccg acaagcagaa gaacggcatc aaggccaact
     3421 tcaagatccg ccacaacatc gaggacggcg gcgtgcagct cgccgaccac taccagcaga
     3481 acacccccat cggcgacggc cccgtgctgc tgcccgacaa ccactacctg agctaccagt
     3541 ccgccctgag caaagacccc aacgagaagc gcgatcacat ggtcctgctg gagttcgtga
     3601 ccgccgccgg gatcactctc ggcatggacg agctgtacaa gtaactagtc gatccaggcc
     3661 tcccagcttt cgtccgtatc atcggtttcg acaacgttcg tcaagttcaa tgcatcagtt
     3721 tcattgccca cacaccagaa tcctactaag tttgagtatt atggcattgg aaaagctgtt
     3781 ttcttctatc atttgttctg cttgtaattt actgtgttct ttcagttttt gttttcggac
     3841 atcaaaatgc aaatggatgg ataagagtta ataaatgata tggtcctttt gttcattctc
     3901 aaattattat tatctgttgt ttttacttta atgggttgaa tttaagtaag aaaggaacta
     3961 acagtgtgat attaaggtgc aatgttagac atataaaaca gtctttcacc tctctttggt
     4021 tatgtcttga attggtttgt ttcttcactt atctgtgtaa tcaagtttac tatgagtcta
     4081 tgatcaagta attatgcaat caagttaagt acagtatagg ctttttgtgt cgaggggcta
     4141 gcgatatcgc ggccgcttcg aaaagcttat aacttcgtat aggatacttt atacgaagtt
     4201 atatgaagac taatcttttt ctctttctca tcttttcact tctcctatca ttatcctcgg
     4261 ccgaattcgg cgccgtgagc aagggagagg aactgttcgg cggcatcgtg cccatcctgg
     4321 tggagctgga gggcgacgtg aacggccaca agttcagcgt gagcggcgag ggcgagggcg
     4381 acgccaccta cggcaagctg accctgaagt tcatctgcac caccggcaag ctgcccgtgc
     4441 cctggcccac cctggtgacc accctgacct ggggcgtgca gtgcttcagc cggtaccccg
     4501 accacatgaa gcagcacgac ttcttcaaga gcgtgatgcc cgagggctac gtgcaggagc
     4561 ggaccatctt cttcaaggac gacggcaact acaagacccg ggccgaggtg aagttcgagg
     4621 gcgacaccct ggtgaaccgg atcgagctga agggcatcga cttcaaggag gacggcaaca
     4681 tcctgggcca caagctggag tacaactaca tcagccacaa cgtgtacatc accgccgaca
     4741 agcagaagaa cggcatcaag gccaacttca aggcccggca caacatcacc gacggcagcg
     4801 tgcagctggc cgaccactac cagcagaaca cccccatcgg cgacggcccc gtgatcctgc
     4861 ccgacaacca ctacctgagc acccagagcg ccctgagcaa ggaccccaac gagaagcggg
     4921 accacatggt gctgctggag ttcgtgaccg ccgccggcat cacccacggc atggacgaac
     4981 tgtacaaaca tgatgagctt taactcgagt agcgatcgtt caaacatttg gcaataaagt
     5041 ttcttaagat tgaatcctgt tgccggtctt gcgatgatta tcatataatt tctgttgaat
     5101 tacgttaagc atgtaataat taacatgtaa tgcatgacgt tatttatgag atgggttttt
     5161 atgattagag tcccgcaatt atacatttaa tacgcgatag aaaacaaaat atagcgcgca
     5221 aactaggata aattatcgcg cgcggtgtca tctatgttac tagatcggga attgatcccc
     5281 cctcgacaga cgcgtataac ttcgtatagt ataccttata cgaagttata tgaagactaa
     5341 tctttttctc tttctcatct tttcacttct cctatcatta tcctcggccg aattcaccgg
     5401 tagcgagctg attaaggaga acatgcacat gaagctgtac atggagggca ccgtgaacaa
     5461 ccaccacttc aagtgcacat ccgagggcga aggcaagccc tacgagggca cccagaccat
     5521 gagaatcaag gtggtcgagg gcggccctct ccccttcgcc ttcgacatcc tggctaccag
     5581 cttcatgtac ggcagcagaa ccttcatcaa ccacacccag ggcatccccg acttctttaa
     5641 gcagtccttc cctgagggct tcacatggga gagagtcacc acatacgaag acgggggcgt
     5701 gctgaccgct acccaggaca ccagcctcca ggacggctgc ctcatctaca acgtcaagat
     5761 cagaggggtg aacttcccat ccaacggccc tgtgatgcag aagaaaacac tcggctggga
     5821 ggccaacacc gagatgctgt accccgctga cggcggcctg gaaggcagaa gcgacatggc
     5881 cctgaagctc gtgggcgggg gccacctgat ctgcaacttc aagaccacat acagatccaa
     5941 gaaacccgct aagaacctca agatgcccgg cgtctactat gtggaccaca gactggaaag
     6001 aatcaaggag gccgacaaag agacctacgt cgagcagcac gaggtggctg tggccagata
     6061 ctgcgacctc cctagcaaac tggggcacaa gcatgatgag ctttaacaat tgggatccgg
     6121 tacaagtaag aattcggtac gctgaaatca ccagtctctc tctacaaatc tatctctctc
     6181 tattttctcc ataaataatg tgtgagtagt ttcccgataa gggaaattag ggttcttata
     6241 gggtttcgct catgtgttga gcatataaga aacccttagt atgtatttgt atttgtaaaa
     6301 tacttctatc aataaaattt ctaattccta aaaccaaaat ccagtactaa aatccagatc
     6361 gatgggcccg gtacccaatt cgccctatag tgagtcgtat tacgcgcgct cactggccgt
     6421 cgttttacaa cgtcgtgact gggaaaaccc tggcgttacc caacttaatc gccttgcagc
     6481 acatccccct ttcgccagct ggcgtaatag cgaagaggcc cgcaccgatc gcccttccca
     6541 acagttgcgc agcctgaatg gcgaatggaa attgtaagcg ttaatgttat cgtaccccta
     6601 ctccaaaaat gtcaaagata cagtctcaga agaccaaagg gctattgaga cttttcaaca
     6661 aagggtaatt tcgggaaacc tcctcggatt ccattgccca gctatctgtc acttcatcga
     6721 aaggacagta gaaaaggaag gtggctccta caaatgccat cattgcgata aaggaaaggc
     6781 tatcattcaa gatgcctctg ccgacagtgg tcccaaagat ggacccccac ccacgaggag
     6841 catcgtggaa aaagaagacg ttccaaccac gtcttcaaag caagtggatt gatgtgacat
     6901 ctccactgac gtaagggatg acgcacaatc ccactatcct tcgcaagacc cttcctctat
     6961 ataaggaagt tcatttcatt tggagaggac agcccaagct gggtacccct agaatttgcc
     7021 atcatggttc gaccattgaa ctgcatcgtc gccgtgtccc aaaatatggg gattggcaag
     7081 aacggagacc gaccctggcc tccgctcagg aacgagttca agtacttcca aagaatgacc
     7141 acaacctctt cagtggaagg taaacagaat ctggtgatta tgggtaggaa aacctggttc
     7201 tccattcctg agaagaatcg acctttaaag gacagaatta atatagttct cagtagagaa
     7261 ctcaaagaac caccacgagg agctcatttt cttgccaaaa gtttggatga tgccttaaga
     7321 cttattgaac aaccggaatt ggcaagtaaa gtagacatgg tttggatagt cggaggcagt
     7381 tctgtttacc aggaagccat gaatcaacca ggccacctca gactctttgt gacaaggatc
     7441 atgcaggaat ttgaaagtga cacgtttttc ccagaaattg atttggggaa atataaactt
     7501 ctcccagaat acccaggcgt cctctctgag gtccaggagg aaaaaggcat caagtataag
     7561 tttgaagtct acgagaagaa agactaacag gaagatgctt tcaagttctc tgctcccctc
     7621 ctaaagctat gcatttttat aagaccatgg gacttttgct ggctttagat ccttcgcggg
     7681 acgtcctttg tttacgtccc gtcggcgctg aatcccccgc tgaaatcacc agtctctctc
     7741 tacaaatcta tctctctcta taataatgtg tgagtagttc ccagataagg gaattagggt
     7801 tcttataggg tttcgctcat gtgttgagca tataagaaac ccttagtatg tatttgtatt
     7861 tgtaaaatac ttctatcaat aaaatttcta attcctaaaa ccaaaatcca ggggtaccga
     7921 gctcgaatta attcggtacg ctgaaatcac cagtctctct ctacaaatct atctctctct
     7981 attttctcca taaataatgt gtgagtagtt tcccgataag ggaaattagg gttcttatag
     8041 ggtttcgctc atgtgttgag catataagaa acccttagta tgtatttgta tttgtaaaat
     8101 acttctatca ataaaatttc taattcctaa aaccaaaatc cagtactaaa atccagatcg
     8161 ataacgttac accacaatat atcctgccaa gatctctaat tccggggatc ggaaatccag
     8221 aagcccgaga ggttgccgcc tttcgggctt tttctttttc aaaaaaaaaa atttataaaa
     8281 cgatctgttg cggccggccg ccgggttgtg ggcaaaggcg ctggcgctcg acggtgggca
     8341 accgcttgcg gttgtccacg ggcggagccg gtgcgcgtag cgcattgtcc acaagccaag
     8401 ggcgaccaat aattgatata tatattcata attgaaaagc taattgaaca tactacttgc
     8461 tgtaactact tgccggagcg aggggtgttt gcaagctgtt gatctgaaag ggctattagc
     8521 gttctcacgt gcctttttga ttagcgattt cacgtgacct tattagcgat ttcacgtact
     8581 ccgattagcg atttcacgta ccctgattag cgatttcacg tggatagttt ttggagcggg
     8641 ccggaaagcc ccgtgaatca aggctttgcg gggcattagc ggtttcacgt ggataactac
     8701 cctctatcca caggcttccg gggataaaaa
//
