LOCUS       sequence-233110-        4750 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Constitutive expression of GFP+ on pAL5000/pMB1 backbone with 
            kanamycin resistance..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4750)
  AUTHORS   DeLorenzo DM, Rottinghaus AG, Henson WR, Moon TS
  TITLE     Molecular Toolkit for Gene Expression Control and Genome 
            Modification in Rhodococcus opacus PD630.
  JOURNAL   ACS Synth Biol. 2018 Feb 16;7(2):727-738. doi: 
            10.1021/acssynbio.7b00416. Epub 2018 Feb 5.
  PUBMED    29366319
REFERENCE   2  (bases 1 to 4750)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1021/acssynbio.7b00416"; journalName: "ACS Synth Biol"; date: 
            "2018-02-16- 16"; volume: "7"; issue: "2"; pages: "727-738"
COMMENT     Sequence Label: pDD57
FEATURES             Location/Qualifiers
     source          1..4750
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      20..47
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     CDS             join(316..318,319..321,322..1035)
                     /codon_start=1
                     /product="Cycle 3 GFP (Crameri et al., 1996)"
                     /label=Cycle 3 GFP
                     /note="mammalian codon-optimized"
                     /translation="MASKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTISFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIK
                     ANFKIRHNMEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITHGMDELYK"
     primer_bind     complement(349..377)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     972..992
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     primer_bind     complement(1060..1077)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1231..1819)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1311..1330)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(2103..2897)
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(2214..2233)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     2824..2843
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
ORIGIN
        1 actagttgat tagctaagca gaaggccatc ctgacggatg gcctttttgc gtttaatact
       61 gtttaaactc tagaaatatt ggatcgtcgg caccgtcacg gccgtgggag gcggcacgat
      121 ccgcgacgtg atgatcggcc gcatccccac ggtgctgcgc agtgagctct acgccatccc
      181 ggcgttgatc tgtgcgttcg ctgtgcgggc tctaacacgt cctagtatgg taggatgagc
      241 aacatttcga cgccgagaga ttcgccgccc gaaatgagca cgatccgcat gcttaattaa
      301 gaaggagata tacatatggc tagcaaagga gaagaacttt tcactggagt tgtcccaatt
      361 cttgttgaat tagatggtga tgttaatggg cacaaatttt ctgtcagtgg agagggtgaa
      421 ggtgatgcta catacggaaa gcttaccctt aaatttattt gcactactgg aaaactacct
      481 gttccatggc caacacttgt cactactttg acctatggtg ttcaatgctt ttcccgttat
      541 ccggatcata tgaaacggca tgactttttc aagagtgcca tgcccgaagg ttatgtacag
      601 gaacgcacta tatctttcaa agatgacggg aactacaaga cgcgtgctga agtcaagttt
      661 gaaggtgata cccttgttaa tcgtatcgag ttaaaaggta ttgattttaa agaagatgga
      721 aacattctcg gacacaaact cgagtacaac tataactcac acaatgtata catcacggca
      781 gacaaacaaa agaatggaat caaagctaac ttcaaaattc gccacaacat ggaagatggc
      841 tccgttcaac tagcagacca ttatcaacaa aatactccaa ttggcgatgg ccctgtcctt
      901 ttaccagaca accattacct gtcgacacaa tctgcccttt cgaaagatcc caacgaaaag
      961 cgtgaccaca tggtccttct tgagtttgta actgctgctg ggattacaca tggcatggat
     1021 gagctctaca aataatgaat tcgagctgga tccagcttcc tcgctcactg actcgctgcg
     1081 ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc
     1141 cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag
     1201 gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca
     1261 tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca
     1321 ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg
     1381 atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag
     1441 gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt
     1501 tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca
     1561 cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg
     1621 cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa gaacagtatt
     1681 tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc
     1741 cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg
     1801 cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg
     1861 gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta
     1921 gatccttttg gggtgggcga agaactccag catgagatcc ccgcgctgga ggatcatcca
     1981 gccggcgtcc cggaaaacga ttccgaagcc caacctttca tagaaggcgg cggtggaatc
     2041 gaaatctcgt gatggcaggt tgggcgtcgc ttggtcggtc atttcgaacc ccagagtccc
     2101 gctcagaaga actcgtcaag aaggcgatag aaggcgatgc gctgcgaatc gggagcggcg
     2161 ataccgtaaa gcacgaggaa gcggtcagcc cattcgccgc caagctcttc agcaatatca
     2221 cgggtagcca acgctatgtc ctgatagcgg tccgccacac ccagccggcc acagtcgatg
     2281 aatccagaaa agcggccatt ttccaccatg atattcggca agcaggcatc gccatgggtc
     2341 acgacgagat cctcgccgtc gggcatccgc gccttgagcc tggcgaacag ttcggctggc
     2401 gcgagcccct gatgctcttc gtccagatca tcctgatcga caagaccggc ttccatccga
     2461 gtacgtgctc gctcgatgcg atgtttcgct tggtggtcga atgggcaggt agccggatca
     2521 agcgtatgca gccgccgcat tgcatcagcc atgatggata ctttctcggc aggagcaagg
     2581 tgagatgaca ggagatcctg ccccggcact tcgcccaata gcagccagtc ccttcccgct
     2641 tcagtgacaa cgtcgagcac agctgcgcaa ggaacgcccg tcgtggccag ccacgatagc
     2701 cgcgctgcct cgtcttggag ttcattcagg gcaccggaca ggtcggtctt gacaaaaaga
     2761 accgggcgcc cctgcgctga cagccggaac acggcggcat cagagcagcc gattgtctgt
     2821 tgtgcccagt catagccgaa tagcctctcc acccaagcgg ccggagaacc tgcgtgcaat
     2881 ccatcttgtt caatcatgcg aaacgatcct catcctgtct cttgatcaga tcttgatccc
     2941 ctgcgccatc agatccttgg cggcaagaaa gccatccagt ttactttgca gggcttccca
     3001 accttaccag agggcgcccc agctggcaat tccggttcgc ttgctgtcca taaaaccgcc
     3061 cagtctagct atcgccatgt aagcccactg caagctacct gctttctctt tgcgcttgcg
     3121 tctgacattc atccggggtc agcaccgttt ctgcggactg gctttctacg tgttccgctt
     3181 cctttagcag cccttgcgcc ctgagtgctt gcggcagcgt gaagctagcg tctaccagga
     3241 cttttacctg tccgacccgt tgcaacggaa ccccccacgg aacccccgcg acacccgctc
     3301 cccaattgcg ttagaacagc ggtggattgt cggcttcgtt gtgggccttt tgagccgctt
     3361 cctgttctgc cgcacgctct ttcctcgccc gatagccgag tcgcttaacg gtgtccagat
     3421 gcagcccgaa atgtttggcc gtttgcggcc aagagtggcc ctcgtcgtcg tgataggcgc
     3481 ggatgcgttc gcggcgtgca gcctgctcgg cgagccactc gctgcgttcc tgcgccacga
     3541 gccggacgac gtggcgttcg gatagtccgg tgattcgagc gccttcggcg gcggtcacgc
     3601 gccgcttttt gcggacagtc ggctgccggt tgtagccgtc gctgtagccg tcgctcatag
     3661 caatgcctcc atggctgacg cggactttgc gcgccgcgca actgtgctcg ccgccgtgcg
     3721 cgctgctgcg cccttccgcg agatggccga ctggcgcgca ctgagtgtgg cctcgtagac
     3781 cacgatcccg tccgcccaaa tgcgcgactt ggttgtgatc caacgccaaa tgctgttggc
     3841 gatggcgcgg acctcgctgt ccggtagcgg tccgggacac acgtcgttgc acgggaactc
     3901 ggcgtttcgc gcgtggcact cggcatagat cgcgcggccg agtccgtcca cgttccgggt
     3961 cggcaggtag atccgcatga gggcgggacg ataggcccac aacctgacgg aatcgaacag
     4021 tgcgcaattc cgccctagcg gcgtcggagc cgctttgtac gtggtctgct gacgccagcg
     4081 cggcggtggc atgttcgcgc cgagctcggc ctcgatgtgg ctgagtgtgt agagatctga
     4141 gtggagccat tccgtttccc aggcgatgtg gccggggttt ttggtcatga ggcctgagta
     4201 actgcggtcg ccgtccacgg cgcgccgaag gccttcggcg cacgccgcca tgtatgcgag
     4261 cggcttacgc cgcgcgtatt cggtgcgtgg aacaggggcg ttgagtgccc acactgcgtg
     4321 tgcgtggccg ttggcgcgat tgcccacgat cgcgttgggc agcggatggg acccccgggc
     4381 gctgagcgct cggagcgctg cgtctggatg gtctacgtcc acgaccagca ggtttgccag
     4441 cgctgttggg ttcgcctcga tgtaccggcg gcctagggcc gacgcgcggc tttggcggta
     4501 gatcccctcg agcagatcgt cgcttgccag cggccagtac ggcagccaga gctgctcaaa
     4561 ttcgtcggcg acgtggctca cgcttggtag tagaccacga ttaatcaccg gtgtatggtc
     4621 cgacacgagc tccaagtcag atatttcgct gaggggccac cccacaactg cacactcccc
     4681 cgctctcccg tcgagccctg gtggtggaac accagcgaca gccgagcacc cccaaccacc
     4741 tgtaccaacc
//
