LOCUS       sequence-128087-        3962 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  TOPO construct expressing ZeoR for generation of GMAP-compatible 
            gene.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3962)
  AUTHORS   Akama-Garren EH, Joshi NS, Tammela T, Chang GP, Wagner BL, Lee DY, 
            Rideout Iii WM, Papagiannakopoulos T, Xue W, Jacks T
  TITLE     A Modular Assembly Platform for Rapid Generation of DNA Constructs.
  JOURNAL   Sci Rep. 2016 Feb 18;6:16836. doi: 10.1038/srep16836.
  PUBMED    26887506
REFERENCE   2  (bases 1 to 3962)
  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"; journalName: "Sci Rep."; 
            date: "2016-02-18"; volume: "6:16836. doi"; pages: " 
            10.1038/srep16836"
COMMENT     Sequence Label: TOPO ZeoR
FEATURES             Location/Qualifiers
     source          1..3962
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    107..128
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(143..148,149..166,167..173)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    181..197
                     /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     186..208
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     205..221
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     205..221
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        239..257
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     239..256
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     CDS             332..706
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     primer_bind     complement(849..868)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(850..868)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(875..892)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(875..891)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(884..906)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     CDS             1029..1331
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="QFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDKV
                     SRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     1210..1229
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             1680..2474
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             1680..2474
                     /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
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(1734..1753)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2344..2363
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     regulatory      2675..2684
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2681..3055
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     rep_origin      3193..3781
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3682..3701
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3935..3952
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 agcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc
       61 acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc
      121 tcactcatta ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa
      181 ttgtgagcgg ataacaattt cacacaggaa acagctatga ccatgattac gccaagctat
      241 ttaggtgaca ctatagaata ctcaagctat gcatcaagct tggtaccgag ctcggatcct
      301 aactcgaacg ctagctgtgc gatcgtttac catggccaag ttgaccagtg ccgttccggt
      361 gctcaccgcg cgcgacgtcg ccggagcggt cgagttctgg accgaccggc tcgggttctc
      421 ccgggacttc gtggaggacg acttcgccgg tgtggtccgg gacgacgtga ccctgttcat
      481 cagcgcggtc caggaccagg tggtgccgga caacaccctg gcctgggtgt gggtgcgcgg
      541 cctggacgag ctgtacgccg agtggtcgga ggtcgtgtcc acgaacttcc gggacgcctc
      601 cgggccggcc atgaccgaga tcggcgagca gccgtggggg cgggagttcg ccctgcgcga
      661 cccggccggc aactgcgtgc acttcgtggc cgaggagcag gactgaaaac gttgttgttt
      721 ggggttgaat tactctggat ccactagtaa cggccgccag tgtgctggaa ttcgccctta
      781 agggcgaatt ctgcagatat ccatcacact ggcggccgct cgagcatgca tctagagggc
      841 ccaattcgcc ctatagtgag tcgtattaca attcactggc cgtcgtttta caacgtcgtg
      901 actgggaaaa ccctggcgtt acccaactta atcgccttgc agcacatccc cctttcgcca
      961 gctggcgtaa tagcgaagag gcccgcaccg atcgcccttc ccaacagttg cgcagcctat
     1021 acgtacggca gtttaaggtt tacacctata aaagagagag ccgttatcgt ctgtttgtgg
     1081 atgtacagag tgatattatt gacacgccgg ggcgacggat ggtgatcccc ctggccagtg
     1141 cacgtctgct gtcagataaa gtctcccgtg aactttaccc ggtggtgcat atcggggatg
     1201 aaagctggcg catgatgacc accgatatgg ccagtgtgcc ggtctccgtt atcggggaag
     1261 aagtggctga tctcagccac cgcgaaaatg acatcaaaaa cgccattaac ctgatgttct
     1321 ggggaatata aatgtcaggc atgagattat caaaaaggat cttcacctag atccttttca
     1381 cgtagaaagc cagtccgcag aaacggtgct gaccccggat gaatgtcagc tactgggcta
     1441 tctggacaag ggaaaacgca agcgcaaaga gaaagcaggt agcttgcagt gggcttacat
     1501 ggcgatagct agactgggcg gttttatgga cagcaagcga accggaattg ccagctgggg
     1561 cgccctctgg taaggttggg aagccctgca aagtaaactg gatggctttc ttgccgccaa
     1621 ggatctgatg gcgcagggga tcaagctctg atcaagagac aggatgagga tcgtttcgca
     1681 tgattgaaca agatggattg cacgcaggtt ctccggccgc ttgggtggag aggctattcg
     1741 gctatgactg ggcacaacag acaatcggct gctctgatgc cgccgtgttc cggctgtcag
     1801 cgcaggggcg cccggttctt tttgtcaaga ccgacctgtc cggtgccctg aatgaactgc
     1861 aagacgaggc agcgcggcta tcgtggctgg ccacgacggg cgttccttgc gcagctgtgc
     1921 tcgacgttgt cactgaagcg ggaagggact ggctgctatt gggcgaagtg ccggggcagg
     1981 atctcctgtc atctcacctt gctcctgccg agaaagtatc catcatggct gatgcaatgc
     2041 ggcggctgca tacgcttgat ccggctacct gcccattcga ccaccaagcg aaacatcgca
     2101 tcgagcgagc acgtactcgg atggaagccg gtcttgtcga tcaggatgat ctggacgaag
     2161 agcatcaggg gctcgcgcca gccgaactgt tcgccaggct caaggcgagc atgcccgacg
     2221 gcgaggatct cgtcgtgacc catggcgatg cctgcttgcc gaatatcatg gtggaaaatg
     2281 gccgcttttc tggattcatc gactgtggcc ggctgggtgt ggcggaccgc tatcaggaca
     2341 tagcgttggc tacccgtgat attgctgaag agcttggcgg cgaatgggct gaccgcttcc
     2401 tcgtgcttta cggtatcgcc gctcccgatt cgcagcgcat cgccttctat cgccttcttg
     2461 acgagttctt ctgaattatt aacgcttaca atttcctgat gcggtatttt ctccttacgc
     2521 atctgtgcgg tatttcacac cgcatacagg tggcactttt cggggaaatg tgcgcggaac
     2581 ccctatttgt ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc
     2641 ctgataaatg cttcaataat agcacgtgag gagggccacc atggccaagt tgaccagtgc
     2701 cgttccggtg ctcaccgcgc gcgacgtcgc cggagcggtc gagttctgga ccgaccggct
     2761 cgggttctcc cgggacttcg tggaggacga cttcgccggt gtggtccggg acgacgtgac
     2821 cctgttcatc agcgcggtcc aggaccaggt ggtgccggac aacaccctgg cctgggtgtg
     2881 ggtgcgcggc ctggacgagc tgtacgccga gtggtcggag gtcgtgtcca cgaacttccg
     2941 ggacgcctcc gggccggcca tgaccgagat cggcgagcag ccgtgggggc gggagttcgc
     3001 cctgcgcgac ccggccggca actgcgtgca cttcgtggcc gaggagcagg actgacacgt
     3061 gctaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg ataatctcat
     3121 gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg tagaaaagat
     3181 caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa
     3241 accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa
     3301 ggtaactggc ttcagcagag cgcagatacc aaatactgtc cttctagtgt agccgtagtt
     3361 aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt
     3421 accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata
     3481 gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt
     3541 ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac
     3601 gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga
     3661 gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg
     3721 ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa
     3781 aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat
     3841 gttctttcct gcgttatccc ctgattctgt ggataaccgt attaccgcct ttgagtgagc
     3901 tgataccgct cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg aggaagcgga
     3961 ag
//
