LOCUS       Exported                5440 bp DNA     circular SYN 23-MAR-2022
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  recombinant plasmid
REFERENCE   1  (bases 1 to 5440)
  AUTHORS   EJPARKER
  TITLE     Direct Submission
  JOURNAL   Exported Apr 27, 2022 from SnapGene 6.0.2
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..5440
                     /mol_type="other DNA"
                     /organism="recombinant plasmid"
     repeat_region   274..302
                     /label=IR1
     rep_origin      587..975
                     /label=R6K gamma ori
                     /note="gamma replication origin from E. coli plasmid R6K; 
                     requires the R6K initiator protein pi for replication"
     primer_bind     complement(1350..1377)
                     /label=pHimarEm1+XbaI_RV2
     primer_bind     1357..1399
                     /label=Fragment.FOR
     primer_bind     1380..1405
                     /locus_tag=""
                     /label=Geneious name: MCS primer
                     /note="Geneious name: MCS primer"
     primer_bind     1380..1405
                     /label=pFru97 MCS fw
     primer_bind     1380..1403
                     /label=cassette_pAG408overhang_FWD
     primer_bind     1380..1400
                     /label=REV_pFru_ins
     primer_bind     1380..1399
                     /label=cassette5_pGRG36overhang_FWD
     primer_bind     1390..1426
                     /label=FWD_primer_PnptII
     promoter        1417..1753
                     /locus_tag=""
                     /label=p-nptII promoter
                     /label=p-nptII
     primer_bind     1417..1436
                     /label=11x-nptII-FL-fw
     primer_bind     1417..1431
                     /label=lederm_nptII.fw
     primer_bind     complement(1566..1590)
                     /label=pntpII_1_REV
     primer_bind     1566..1590
                     /label=pntpII_1_FWD
     primer_bind     1567..1585
                     /label=pFru97xpMRE100_nptIIGib_rv
     primer_bind     complement(1688..1708)
                     /label=pntpII_2_REV
     primer_bind     1688..1708
                     /label=pntpII_2_FWD
     primer_bind     complement(1732..1766)
                     /label=rbs_1_REV
     primer_bind     1732..1766
                     /label=rbs_1_FWD
     primer_bind     complement(1752..1781)
                     /label=REV_primer_PnptII_mTq2/sG2
     primer_bind     complement(1752..1774)
                     /label=REV_primer_PnptII_mCl/mO/mSc/sY2
     primer_bind     complement(1752..1774)
                     /label=REV_primer_PnptII_mTagBFP2
     primer_bind     1762..1791
                     /label=FWD_primer_mTq2/sGFP2
     primer_bind     1762..1788
                     /label=FWD_primer_mCl/mO/mSc/sY2
     CDS             1772..2489
                     /codon_start=1
                     /locus_tag=""
                     /product=""
                     /label=sYFP2
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIKA
                     NFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSKLSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     1773..1788
                     /label=FWD_sYFP2
     primer_bind     1773..1786
                     /label=XFPs for
     primer_bind     1773..1784
                     /label=XhoI_Ptac_XFP_for
     primer_bind     complement(2290..2310)
                     /label=FWD_primer_PnptII
     primer_bind     complement(2436..2455)
                     /label=REV_primer_sG2
     primer_bind     complement(2466..2509)
                     /label=REV_primer_mTq2
     primer_bind     complement(2467..2489)
                     /label=nextgenFP.rv
     primer_bind     complement(2467..2488)
                     /label=NotI_XFP_rev
     primer_bind     complement(2469..2509)
                     /label=REV_primer_mO
     primer_bind     complement(2469..2488)
                     /label=11x-FL-rv
     primer_bind     complement(2470..2509)
                     /label=REV_primer_sY2
     primer_bind     complement(2470..2489)
                     /label=REV_sYFP2
     primer_bind     complement(2470..2489)
                     /label=Ptac+eXFP_rev
     primer_bind     complement(2471..2489)
                     /label=XFPs rev KpnI
     primer_bind     complement(2497..2516)
                     /label=FWD_pFru_ins
     primer_bind     2500..2522
                     /label=Gib_Gent.for.cor
     terminator      2506..2600
                     /locus_tag=""
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             2644..3303
                     /codon_start=1
                     /locus_tag=""
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(2705..2733)
                     /label=pMRE130inserts_rev
     primer_bind     complement(3288..3303)
                     /label=Chloramp_rev
     terminator      3368..3454
                     /locus_tag=""
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB ge
                     ne"
     primer_bind     complement(3459..3476)
                     /label=Gib_Gent.for
     primer_bind     complement(3464..3503)
                     /label=Fragment.REV
     primer_bind     complement(3464..3481)
                     /label=cassette_pGRG36overhang_RV
     primer_bind     complement(3464..3481)
                     /label=pMRE130_cassette2_RV
     primer_bind     complement(3465..3481)
                     /label=cassette_pAG408overhang_RV
     primer_bind     3482..3508
                     /label=pHimarEm1+XmaI_FWD
     repeat_region   3571..3599
                     /label=IR2
     primer_bind     3845..3861
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3941..3952)
                     /label=cassette_pAG408overhang_RV
     oriT            3948..4056
                     /note="incP origin of transfer"
     protein_bind    4203..4224
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        4239..4269
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     promoter        4239..4268
                     /gene="lac prom"
                     /label=lac prom promoter
     protein_bind    4277..4293
                     /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)."
     misc_feature    4313..5359
                     /label=Transposase
                     /label=nonstandard type:  ORF
                     /note="/sequence=ORF_2 rf(1)"
ORIGIN
        1 ctagatgcat gctcgagcgg ccgccagtgt gatggatttg aaaaaggaag agtatgagta
       61 ttcaacattt ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg
      121 ctcacccaga aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg
      181 gttacatcga actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac
      241 gttttccaat gatgagcact tttaaagttc tgctaacagg ttggctgata agtccccggt
      301 ctgacacata gatggcgtcg ctagtattaa atgcatatta tttttatata gtaccaacct
      361 tcaaatgatt cccagatctc aaactggaac aacactcaac cctatctcgg tctattcttt
      421 tgatttataa gggattttgc cgatttcggc ctattggtta aaaaatgagc tgatttaaca
      481 aaaatttaac gcgaatttta acaaaatatt aacgcttaca atttaggtgg cacttttcgg
      541 ggaaatgtgc gcggaaccag atccccacga tgcgtccggc gtagaggatc tgaagatcag
      601 cagttcaacc tgttgatagt acgtactaag ctctcatgtt tcacgtacta agctctcatg
      661 tttaacgtac taagctctca tgtttaacga actaaaccct catggctaac gtactaagct
      721 ctcatggcta acgtactaag ctctcatgtt tcacgtacta agctctcatg tttgaacaat
      781 aaaattaata taaatcagca acttaaatag cctctaaggt tttaagtttt ataagaaaaa
      841 aaagaatata taaggctttt aaagctttta aggtttaacg gttgtggaca acaagccagg
      901 gatgtaacgc actgagaagc ccttagagcc tctcaaagca attttcagtg acacaggaac
      961 acttaacggc tgacagaatt cttgaagacg aaagggcctc gtgatacgcc tatttttata
     1021 ggttaatgtc atgataataa tggtttctta gacgtcaggt ggcacttttc ggggttaatt
     1081 aagggctgca ggaattcgat atcaagctta tcgataccgt cgaaattctg attaataatt
     1141 tgtttaaatt tttcgtttgg cgtgaggtat ccaagtcttt tacgaggtcg attattgagt
     1201 ttattttcaa tccacttaat ctgtttgttg gttacttcac taaagtcctt accctttggg
     1261 atatactgcc tgataagccc gttggtgttt tcattggcac cacgttccca tgagtggtat
     1321 ggtttgcaaa aatagaattt tatttccaat ttttgcgcaa tttcctcgtg ctctagaata
     1381 taaactgcca ggaattgggg atcggattcg agctcgcacg ctgccgcaag cactcagggc
     1441 gcaagggctg ctaaaggaag cggaacacgt agaaagccag tccgcagaaa cggtgctgac
     1501 cccggatgaa tgtcagctac tgggctatct ggacaaggga aaacgcaagc gcaaagagaa
     1561 agcaggtagc ttgcagtggg cttacatggc gatagctaga ctgggcggtt ttatggacag
     1621 caagcgaacc ggaattgcca gctggggcgc cctctggtaa ggttgggaag ccctgcaaag
     1681 taaactggat ggctttcttg ccgccaagga tctgatggcg caggggatca agatctgatc
     1741 aagagacagg atactagtgg aggaagaaaa aatgagcaag ggcgaggagc tgttcaccgg
     1801 ggtggtgccc atcctggtcg agctggacgg cgacgtaaac ggccacaagt tcagcgtgtc
     1861 cggcgagggc gagggcgatg ccacctacgg caagctgacc ctgaagttca tctgcaccac
     1921 cggcaagctg cccgtgccct ggcccaccct cgtgaccacc ctgacctacg gcgtgcagtg
     1981 cttcgcccgc taccccgacc acatgaagca gcacgacttc ttcaagtccg ccatgcccga
     2041 aggctacgtc caggagcgca ccatcttctt caaggacgac ggcaactaca agacccgcgc
     2101 cgaggtgaag ttcgagggcg acaccctggt gaaccgcatc gagctgaagg gcatcgactt
     2161 caaggaggac ggcaacatcc tggggcacaa gctggagtac aactacaaca gccacaacgt
     2221 ctatatcacc gccgacaagc agaagaacgg catcaaggcc aacttcaaga tccgccacaa
     2281 catcgaggac ggcggcgtgc agctcgccga ccactaccag cagaacaccc ccatcggcga
     2341 cggccccgtg ctgctgcccg acaaccacta cctgagctac cagtccaagc tgagcaaaga
     2401 ccccaacgag aagcgcgatc acatggtcct gctggagttc gtgaccgccg ccgggatcac
     2461 tctcggcatg gacgagctgt acaagtaagt aattagctga gcttggactc ctgttgatag
     2521 atccagtaat gacctcagaa ctccatctgg atttgttcag aacgctcggt tgccgccggg
     2581 cgttttttat tggtgagaat ccaagctagc ttggcgagat tttcaggagc taaggaagct
     2641 aaaatggaga aaaaaatcac tggatatacc accgttgata tatcccaatg gcatcgtaaa
     2701 gaacattttg aggcatttca gtcagttgct caatgtacct ataaccagac cgttcagctg
     2761 gatattacgg cctttttaaa gaccgtaaag aaaaataagc acaagtttta tccggccttt
     2821 attcacattc ttgcccgcct gatgaatgct catccggaat ttcgtatggc aatgaaagac
     2881 ggtgagctgg tgatatggga tagtgttcac ccttgttaca ccgttttcca tgagcaaact
     2941 gaaacgtttt catcgctctg gagtgaatac cacgacgatt tccggcagtt tctacacata
     3001 tattcgcaag atgtggcgtg ttacggtgaa aacctggcct atttccctaa agggtttatt
     3061 gagaatatgt ttttcgtctc agccaatccc tgggtgagtt tcaccagttt tgatttaaac
     3121 gtggccaata tggacaactt cttcgccccc gttttcacca tgggcaaata ttatacgcaa
     3181 ggcgacaagg tgctgatgcc gctggcgatt caggttcatc atgccgtctg tgatggcttc
     3241 catgtcggca gaatgcttaa tgaattacaa cagtactgcg atgagtggca gggcggggcg
     3301 taattttttt aaggcagtta ttggtgccct taaacgcctg gggtaatgac tctctagctt
     3361 gaggcatcaa ataaaacgaa aggctcagtc gaaagactgg gcctttcgtt ttatctgttg
     3421 tttgtcggtg aacgctctcc tgagtaggac aaatccgccg ctaggagctt gcggccgcca
     3481 gcccgggcaa ttcgaggggt atcgctcttg aagggaacta tgttgaataa taaaaacgaa
     3541 ttttgacaaa aaaatgtgtt tttctttgtt agaccgggga cttatcagcc aacctgttat
     3601 gtggcgcggt attatcccgt attgacgccg ggcaagagca actcggtcgc cgcatacact
     3661 attctcagaa tgacttggtt gagtactcac cagtcacaga aaagcatctt acggatggca
     3721 tgacagtaag agaattatgc agtgctgcca taaccatgag tgataacact gcggccaact
     3781 tacttctgac aacgatcgag gcgattaagt tgggtaacgc cagggttttc ccagtcacga
     3841 cgttgtaaaa cgacggccag tgaattaatt cttgaagacg aaagggcctc gtgatacgcc
     3901 tatttttata ggttaatgtc atgataataa tggtttctta gagcttacgg ccagcctcgc
     3961 agagcaggat tcccgttgag caccgccagg tgcgaataag ggacagtgaa gaaggaacac
     4021 ccgctcgcgg gtgggcctac ttcacctatc ctgcccggct gacgccgttg gatacaccaa
     4081 ggaaagtcta cacgaaccct ttggcaaaat cctgtatatc gtgcgaaaaa ggatggatat
     4141 accgaaaaaa tcgctataat gaccccgaag cagggttatg cagcgggtga gcgcaacgca
     4201 attaatgtga gttagctcac tcattaggca ccccaggctt tacactttat gcttccggct
     4261 cgtatgttgt gtggaattgt gagcggataa caatttcaca caggaaacac atatggaaaa
     4321 aaaggaattt cgtgttttga taaaatactg ttttctgaag ggaaaaaata cggtggaagc
     4381 aaaaacttgg cttgataatg agtttccgga ctctgcccca gggaaatcaa caataattga
     4441 ttggtatgca aaattcaagc gtggtgaaat gagcacggag gacggtgaac gcagtggacg
     4501 cccgaaagag gtggttaccg acgaaaacat caaaaaaatc cacaaaatga ttttgaatga
     4561 ccgtaaaatg aagttgatcg agatagcaga ggccttaaag atatcaaagg aacgtgttgg
     4621 tcatatcatt catcaatatt tggatatgcg gaagctctgt gcaaaatggg tgccgcgcga
     4681 gctcacattt gaccaaaaac aacaacgtgt tgatgattct gagcggtgtt tgcagctgtt
     4741 aactcgtaat acacccgagt ttttccgtcg atatgtgaca atggatgaaa catggctcca
     4801 tcactacact cctgagtcca atcgacagtc ggctgagtgg acagcgaccg gtgaaccgtc
     4861 tccgaagcgt ggaaagactc aaaagtccgc tggcaaagta atggcctctg ttttttggga
     4921 tgcgcatgga ataattttta tcgattatct tgagaaggga aaaaccatca acagtgacta
     4981 ttatatggcg ttattggagc gtttgaaggt cgaaatcgcg gcaaaacggc cccacatgaa
     5041 gaagaaaaaa gtgttgttcc accaagacaa cgcaccgtgc cacaagtcat tgagaacgat
     5101 ggcaaaaatt catgaattgg gcttcgaatt gcttccccac ccaccgtatt ctccagatct
     5161 ggcccccagc gactttttct tgttctcaga cctcaaaagg atgctcgcag ggaaaaaatt
     5221 tggctgcaat gaagaggtga tcgccgaaac tgaggcctat tttgaggcaa aaccgaagga
     5281 gtactaccaa aatggtatca aaaaattgga aggtcgttat aatcgttgta tcgctcttga
     5341 agggaactat gttgaataat aaaaacgaat tttgacaaaa aaatgtgttt ttctttgtta
     5401 gaccggaatc tgcagaattc cagcacactg gcggccgtta
//
