LOCUS       Exported                6973 bp ds-DNA     circular SYN 13-MAY-2021
DEFINITION  Acceptor plasmid for odd level position 2, lacZ blue-white 
            screening. Part of the OpenPlant toolkit..
ACCESSION   .
VERSION     .
KEYWORDS    pCk2
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6973)
  AUTHORS   Sauret-Gueto S, Frangedakis E, Silvestri L, Rebmann M, Tomaselli M, 
            Markel K, Delmans M, West A, Patron NJ, Haseloff J
  TITLE     Systematic tools for reprogramming plant gene expression in a simple
            model, Marchantia polymorpha.
  JOURNAL   ACS Synth Biol. 2020 Mar 12. doi: 10.1021/acssynbio.9b00511.
  PUBMED    32163700
REFERENCE   2  (bases 1 to 6973)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 13, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6973
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     misc_feature    433..457
                     /label=LB T-DNA repeat
                     /note="left border repeat from nopaline C58 T-DNA"
     primer_bind     797..819
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     811..828
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     812..828
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_feature    829..885
                     /label=MCS
                     /note="pUC18/19 multiple cloning site"
     primer_bind     complement(898..914)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(898..914)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(911..933)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    922..938
                     /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)."
     promoter        complement(946..976)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    991..1012
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     misc_feature    1096..1120
                     /label=RB T-DNA repeat
                     /note="right border repeat from nopaline C58 T-DNA"
     CDS             2421..3050
                     /codon_start=1
                     /product="stability protein from the Pseudomonas plasmid 
                     pVS1 (Heeb et al., 2000)"
                     /label=pVS1 StaA
                     /translation="MKVIAVLNQKGGSGKTTIATHLARALQLAGADVLLVDSDPQGSAR
                     DWAAVREDQPLTVVGIDRPTIDRDVKAIGRRDFVVIDGAPQAADLAVSAIKAADFVLIP
                     VQPSPYDIWATADLVELVKQRIEVTDGRLQAAFVVSRAIKGTRIGGEVAEALAGYELPI
                     LESRITQRVSYPGTAAAGTTVLESEPEGDAAREVQALAAEIKSKLI"
     CDS             3479..4552
                     /codon_start=1
                     /product="replication protein from the Pseudomonas plasmid 
                     pVS1 (Heeb et al., 2000)"
                     /label=pVS1 RepA
                     /translation="MSGRKPSGPVQIGAALGDDLVEKLKAAQAAQRQRIEAEARPGESW
                     QAAADRIRKESRQPPAAGAPSIRKPPKGDEQPDFFVPMLYDVGTRDSRSIMDVAVFRLS
                     KRDRRAGEVIRYELPDGHVEVSAGPAGMASVWDYDLVLMAVSHLTESMNRYREGKGDKP
                     GRVFRPHVADVLKFCRRADGGKQKDDLVETCIRLNTTHVAMQRTKKAKNGRLVTVSEGE
                     ALISRYKIVKSETGRPEYIEIELADWMYREITEGKNPDVLTVHPDYFLIDPGIGRFLYR
                     LARRAAGKAEARWLFKTIYERSGSAGEFKKFCFTVRKLIGSNDLPEYDLKEEAGQAGPI
                     LVMRYRNLIEGEASAGS"
     rep_origin      4618..4812
                     /label=pVS1 oriV
                     /note="origin of replication for the Pseudomonas plasmid 
                     pVS1 (Heeb et al., 2000)"
     primer_bind     complement(5048..5070)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    5156..5296
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(5311..5328)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5482..6070)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5562..5581)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(6158..6973)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     6881..6900
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
ORIGIN
        1 aacagaactt attatttcct tcctcttttc tacagtattt aaagataccc caagaagcta
       61 attataacaa gacgaactcc aattcactgt tccttgcatt ctaaaacctt aaataccaga
      121 aaacagcttt ttcaaagttg ttttcaaagt tggcgtataa catagtatcg acggagccga
      181 ttttgaaacc gcggtgatca caggcagcaa cgctctgtca tcgttacaat caacatgcta
      241 ccctccgcga gatcatccgt gtttcaaacc cggcagctta gttgccgttc ttccgaatag
      301 catcggtaac atgagcaaag tctgccgcct tacaacggct ctcccgctga cgccgtcccg
      361 gactgatggg ctgcctgtat cgagtggtga ttttgtgccg agctgccggt cggggagctg
      421 ttggctggct ggtggcagga tatattgtgg tgtaaacata acgaattgct cttcagcaag
      481 gagagagacc cacagcttgt ctgtaagcgg atgccgggag cagacaagcc cgtcagggcg
      541 cgtcagcggg tgttggcggg tgtcggggct ggcttaacta tgcggcatca gagcagattg
      601 tactgagagt gcaccatatg cggtgtgaaa taccgcacag atgcgtaagg agaaaatacc
      661 gcatcaggcg ctcatccgcc attcaggctg cgcaactgtt gggaagggcg atcggcgcag
      721 gcctcttcgc tattacgcca gctggcgaaa gggggatgtg ctgcaaggcg attaagttgg
      781 gtaacgccag ggttttccca gtcacgacgt tgtaaaacga cggccagtga attcgagctc
      841 ggtacccggg gatcctctag agtcgacctg caggcatgca agcttggcgt aatcatggtc
      901 atagctgttt cctgtgtgaa attgttatcc gctcacaatt ccacacaaca tacgagccgg
      961 aagcatatag tgtaaagcct ggggtgccta atgagtgagc taactcacat taattgcgtt
     1021 gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctggggt ctctcgctat
     1081 actgaagagc aattctgaca ggatatattg gcgggtaaac ctaagagaaa agagcgttta
     1141 ttagaataat cggatattta aaagggcgtg aaaaggttta tccgttcgtc catttgtatg
     1201 tgcatgccaa ccacagggtt cccctcggga tcaaagtact ttgatccaac ccctccgctg
     1261 ctatagtgca gtcggcttct gacgttcagt gcagccgtca tctgaaaacg acatgtcgca
     1321 caagtcctaa gttacgcgac aggctgccgc cctgcccttt tcctggcgtt ttcttgtcgc
     1381 gtgttttagt cgcataaagt agaatacttg cgactagaac cggagacatt acgccatgaa
     1441 caagagcgcc gccgctggcc tgctgggcta tgcccgcgtc agcaccgacg accaggactt
     1501 gaccaaccaa cgggccgaac tgcacgcggc cggctgcacc aagctgtttt ccgagaagat
     1561 caccggcacc aggcgcgacc gcccggagct ggccaggatg cttgaccacc tacgccctgg
     1621 cgacgttgtg acagtgacca ggctagaccg cctggcccgc agcacccgcg acctactgga
     1681 cattgccgag cgcatccagg aggccggcgc gggcctgcgt agcctggcag agccgtgggc
     1741 cgacaccacc acgccggccg gccgcatggt gttgaccgtg ttcgccggca ttgccgagtt
     1801 cgagcgttcc ctaatcatcg accgcacccg gagcgggcgc gaggccgcca aggcccgagg
     1861 cgtgaagttt ggcccccgcc ctaccctcac cccggcacag atcgcgcacg cccgcgagct
     1921 gatcgaccag gaaggccgca ccgtgaaaga ggcggctgca ctgcttggcg tgcatcgctc
     1981 gaccctgtac cgcgcacttg agcgcagcga ggaagtgacg cccaccgagg ccaggcggcg
     2041 cggtgccttc cgtgaggacg cattgaccga ggccgacgcc ctggcggccg ccgagaatga
     2101 acgccaagag gaacaagcat gaaaccgcac caggacggcc aggacgaacc gtttttcatt
     2161 accgaagaga tcgaggcgga gatgatcgcg gccgggtacg tgttcgagcc gcccgcgcac
     2221 ctctcaaccg tgcggctgca tgaaatcctg gccggtttgt ctgatgccaa gctggcggcc
     2281 tggccggcca gcttggccgc tgaagaaacc gagcgccgcc gtctaaaaag gtgatgtgta
     2341 tttgagtaaa acagcttgcg tcatgcggtc gctgcgtata tgatgcgatg agtaaataaa
     2401 caaatacgca aggggaacgc atgaaggtta tcgctgtact taaccagaaa ggcgggtcag
     2461 gcaagacgac catcgcaacc catctagccc gcgccctgca actcgccggg gccgatgttc
     2521 tgttagtcga ttccgatccc cagggcagtg cccgcgattg ggcggccgtg cgggaagatc
     2581 aaccgctaac cgttgtcggc atcgaccgcc cgacgattga ccgcgacgtg aaggccatcg
     2641 gccggcgcga cttcgtagtg atcgacggag cgccccaggc ggcggacttg gctgtgtccg
     2701 cgatcaaggc agccgacttc gtgctgattc cggtgcagcc aagcccttac gacatatggg
     2761 ccaccgccga cctggtggag ctggttaagc agcgcattga ggtcacggat ggaaggctac
     2821 aagcggcctt tgtcgtgtcg cgggcgatca aaggcacgcg catcggcggt gaggttgccg
     2881 aggcgctggc cgggtacgag ctgcccattc ttgagtcccg tatcacgcag cgcgtgagct
     2941 acccaggcac tgccgccgcc ggcacaaccg ttcttgaatc agaacccgag ggcgacgctg
     3001 cccgcgaggt ccaggcgctg gccgctgaaa ttaaatcaaa actcatttga gttaatgagg
     3061 taaagagaaa atgagcaaaa gcacaaacac gctaagtgcc ggccgtccga gcgcacgcag
     3121 cagcaaggct gcaacgttgg ccagcctggc agacacgcca gccatgaagc gggtcaactt
     3181 tcagttgccg gcggaggatc acaccaagct gaagatgtac gcggtacgcc aaggcaagac
     3241 cattaccgag ctgctatctg aatacatcgc gcagctacca gagtaaatga gcaaatgaat
     3301 aaatgagtag atgaatttta gcggctaaag gaggcggcat ggaaaatcaa gaacaaccag
     3361 gcaccgacgc cgtggaatgc cccatgtgtg gaggaacggg cggttggcca ggcgtaagcg
     3421 gctgggttgc ctgccggccc tgcaatggca ctggaacccc caagcccgag gaatcggcgt
     3481 gagcggtcgc aaaccatccg gcccggtaca aatcggcgcg gcgctgggtg atgacctggt
     3541 ggagaagttg aaggccgcgc aggccgccca gcggcaacgc atcgaggcag aagcacgccc
     3601 cggtgaatcg tggcaagcgg ccgctgatcg aatccgcaaa gaatcccggc aaccgccggc
     3661 agccggtgcg ccgtcgatta ggaagccgcc caagggcgac gagcaaccag attttttcgt
     3721 tccgatgctc tatgacgtgg gcacccgcga tagtcgcagc atcatggacg tggccgtttt
     3781 ccgtctgtcg aagcgtgacc gacgagctgg cgaggtgatc cgctacgagc ttccagacgg
     3841 gcacgtagag gtttccgcag ggccggccgg catggccagt gtgtgggatt acgacctggt
     3901 actgatggcg gtttcccatc taaccgaatc catgaaccga taccgggaag ggaagggaga
     3961 caagcccggc cgcgtgttcc gtccacacgt tgcggacgta ctcaagttct gccggcgagc
     4021 cgatggcgga aagcagaaag acgacctggt agaaacctgc attcggttaa acaccacgca
     4081 cgttgccatg cagcgtacga agaaggccaa gaacggccgc ctggtgacgg tatccgaggg
     4141 tgaagccttg attagccgct acaagatcgt aaagagcgaa accgggcggc cggagtacat
     4201 cgagatcgag ctagctgatt ggatgtaccg cgagatcaca gaaggcaaga acccggacgt
     4261 gctgacggtt caccccgatt actttttgat cgatcccggc atcggccgtt ttctctaccg
     4321 cctggcacgc cgcgccgcag gcaaggcaga agccagatgg ttgttcaaga cgatctacga
     4381 acgcagtggc agcgccggag agttcaagaa gttctgtttc accgtgcgca agctgatcgg
     4441 gtcaaatgac ctgccggagt acgatttgaa ggaggaggcg gggcaggctg gcccgatcct
     4501 agtcatgcgc taccgcaacc tgatcgaggg cgaagcatcc gccggttcct aatgtacgga
     4561 gcagatgcta gggcaaattg ccctagcagg ggaaaaaggt cgaaaaggac tctttcctgt
     4621 ggatagcacg tacattggga acccaaagcc gtacattggg aaccggaacc cgtacattgg
     4681 gaacccaaag ccgtacattg ggaaccggtc acacatgtaa gtgactgata taaaagagaa
     4741 aaaaggcgat ttttccgcct aaaactcttt aaaacttatt aaaactctta aaacccgcct
     4801 ggcctgtgca taactgtctg gccagcgcac agccgaagtg ctgcaaaaag cgcctaccct
     4861 tcggtcgctg cgctccctac gccccgccgc ttcgcgtcgg cctatcgcgg ccgctggccg
     4921 ctcaaaaatg gctggcctac ggccaggcaa tctaccaggg cgcggacaag ccgcgccgtc
     4981 gccactcgac cgccggcgcc cacatcaagg caccctgcct cgcgcgtttc ggtgatgacg
     5041 gtgaaaacct ctgacacatg cagctcccgg tgacggtcac agcttgtctg taagcggatg
     5101 ccgggagcag acaagcccgt cagggcgcgt cagcgggtgt tggcgggtgt cggggcgcag
     5161 ccatgaccca gtcacgtagc gatagcggag tgtatactgg cttaactatg cggcatcaga
     5221 gcagattgta ctgagagtgc accatatgcg gtgtgaaata ccgcacagat gcgtaaggag
     5281 aaaataccgc atcaggcgct catccgcttc ctcgctcact gactcgctgc gctcggtcgt
     5341 tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc
     5401 aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa
     5461 aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa
     5521 tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc
     5581 ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc
     5641 cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag
     5701 ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga
     5761 ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc
     5821 gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac
     5881 agagttcttg aagtggtggc ctaactacgg ctacactaga aggacagtat ttggtatctg
     5941 cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca
     6001 aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa
     6061 aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa
     6121 ctcacgttaa gggattttgg tcatgcattc taggtgatta gaaaaactca tcgagcatca
     6181 aatgaaactg caatttattc atatcaggat tatcaatacc atatttttga aaaagccgtt
     6241 tctgtaatga aggagaaaac tcaccgaggc agttccatag gatggcaaga tcctggtatc
     6301 ggtctgcgat tccgactcgt ccaacatcaa tacaacctat taatttcccc tcgtcaaaaa
     6361 taaggttatc aagtgagaaa tcaccatgag tgacgactga atccggtgag aatggcaaaa
     6421 gtttatgcat ttctttccag acttgttcaa caggccagcc attacgctcg tcatcaaaat
     6481 cactcgcatc aaccaaaccg ttattcattc gtgattgcgc ctgagcgagt cgaaatacgc
     6541 gatcgctgtt aaaaggacaa ttacaaacag gaatcgaatg caaccggcgc aggaacactg
     6601 ccagcgcatc aacaatattt tcacctgaat caggatattc ttctaatacc tggaatgctg
     6661 ttttccctgg gatcgcagtg gtgagtaacc atgcatcatc aggagtacgg ataaaatgct
     6721 tgatggtcgg aagaggcata aattccgtca gccagtttag tctgaccatc tcatctgtaa
     6781 catcattggc aacgctacct ttgccatgtt tcagaaacaa ctctggcgca tcgggcttcc
     6841 catacaatcg gtagattgtc gcacctgatt gcccgacatt atcgcgagcc catttatacc
     6901 catataaatc agcatccatg ttggaattta atcgcggcct tgagcaagac gtttcccgtt
     6961 gaatatggct cat
//
