LOCUS       sequence-128119-        4286 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  TOPO construct expressing mT2 for generation of GMAP-compatible 
            gene.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4286)
  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 4286)
  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 mT2
FEATURES             Location/Qualifiers
     source          1..4286
                     /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"
     primer_bind     complement(638..657)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     complement(1173..1192)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1174..1192)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1199..1216)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1199..1215)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1208..1230)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     CDS             1353..1655
                     /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     1534..1553
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             2004..2798
                     /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             2004..2798
                     /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(2058..2077)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2668..2687
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     regulatory      2999..3008
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3005..3379
                     /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      3517..4105
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4006..4025
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4259..4276
                     /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 catggtgtcc aagggcgagg aactgttcac
      361 cggcgtggtg cccatcctgg tggaactgga tggcgacgtg aacggccaca agttcagcgt
      421 gtccggcgag ggcgaaggcg acgccacata cggaaagctg accctgaagt tcatctgcac
      481 caccggcaag ctgcccgtgc cttggcctac cctcgtgacc acactgtctt ggggcgtgca
      541 gtgcttcgcc agataccccg accatatgaa gcagcacgac ttcttcaaga gcgccatgcc
      601 cgagggctac gtgcaggaaa gaaccatctt ctttaaagac gacggcaact acaagaccag
      661 ggccgaagtg aagttcgagg gcgacaccct cgtgaacaga atcgagctga agggcatcga
      721 cttcaaagag gacggcaaca tcctgggcca caagctggag tacaactact tcagcgacaa
      781 cgtgtacatc accgccgaca agcagaagaa cggcatcaag gccaacttca agatccggca
      841 caacatcgag gacggcggcg tgcagctggc cgatcactac cagcagaaca cccctatcgg
      901 cgacggccct gtgctgctgc ctgacaacca ctacctgagc acccagagca agctgagcaa
      961 ggaccccaac gagaagaggg accacatggt gctgctggaa ttcgtgaccg ccgctggcat
     1021 caccctgggc atggacgagc tgtacaagaa acgttgttgt ttggggttga attactctgg
     1081 atccactagt aacggccgcc agtgtgctgg aattctgcag atatccatca cactggcggc
     1141 cgctcgagca tgcatctaga gggcccaatt cgccctatag tgagtcgtat tacaattcac
     1201 tggccgtcgt tttacaacgt cgtgactggg aaaaccctgg cgttacccaa cttaatcgcc
     1261 ttgcagcaca tccccctttc gccagctggc gtaatagcga agaggcccgc accgatcgcc
     1321 cttcccaaca gttgcgcagc ctatacgtac ggcagtttaa ggtttacacc tataaaagag
     1381 agagccgtta tcgtctgttt gtggatgtac agagtgatat tattgacacg ccggggcgac
     1441 ggatggtgat ccccctggcc agtgcacgtc tgctgtcaga taaagtctcc cgtgaacttt
     1501 acccggtggt gcatatcggg gatgaaagct ggcgcatgat gaccaccgat atggccagtg
     1561 tgccggtctc cgttatcggg gaagaagtgg ctgatctcag ccaccgcgaa aatgacatca
     1621 aaaacgccat taacctgatg ttctggggaa tataaatgtc aggcatgaga ttatcaaaaa
     1681 ggatcttcac ctagatcctt ttcacgtaga aagccagtcc gcagaaacgg tgctgacccc
     1741 ggatgaatgt cagctactgg gctatctgga caagggaaaa cgcaagcgca aagagaaagc
     1801 aggtagcttg cagtgggctt acatggcgat agctagactg ggcggtttta tggacagcaa
     1861 gcgaaccgga attgccagct ggggcgccct ctggtaaggt tgggaagccc tgcaaagtaa
     1921 actggatggc tttctcgccg ccaaggatct gatggcgcag gggatcaagc tctgatcaag
     1981 agacaggatg aggatcgttt cgcatgattg aacaagatgg attgcacgca ggttctccgg
     2041 ccgcttgggt ggagaggcta ttcggctatg actgggcaca acagacaatc ggctgctctg
     2101 atgccgccgt gttccggctg tcagcgcagg ggcgcccggt tctttttgtc aagaccgacc
     2161 tgtccggtgc cctgaatgaa ctgcaagacg aggcagcgcg gctatcgtgg ctggccacga
     2221 cgggcgttcc ttgcgcagct gtgctcgacg ttgtcactga agcgggaagg gactggctgc
     2281 tattgggcga agtgccgggg caggatctcc tgtcatctca ccttgctcct gccgagaaag
     2341 tatccatcat ggctgatgca atgcggcggc tgcatacgct tgatccggct acctgcccat
     2401 tcgaccacca agcgaaacat cgcatcgagc gagcacgtac tcggatggaa gccggtcttg
     2461 tcgatcagga tgatctggac gaagagcatc aggggctcgc gccagccgaa ctgttcgcca
     2521 ggctcaaggc gagcatgccc gacggcgagg atctcgtcgt gacccatggc gatgcctgct
     2581 tgccgaatat catggtggaa aatggccgct tttctggatt catcgactgt ggccggctgg
     2641 gtgtggcgga ccgctatcag gacatagcgt tggctacccg tgatattgct gaagagcttg
     2701 gcggcgaatg ggctgaccgc ttcctcgtgc tttacggtat cgccgctccc gattcgcagc
     2761 gcatcgcctt ctatcgcctt cttgacgagt tcttctgaat tattaacgct tacaatttcc
     2821 tgatgcggta ttttctcctt acgcatctgt gcggtatttc acaccgcata caggtggcac
     2881 ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac attcaaatat
     2941 gtatccgctc atgagacaat aaccctgata aatgcttcaa taatagcacg tgaggagggc
     3001 caccatggcc aagttgacca gtgccgttcc ggtgctcacc gcgcgcgacg tcgccggagc
     3061 ggtcgagttc tggaccgacc ggctcgggtt ctcccgggac ttcgtggagg acgacttcgc
     3121 cggtgtggtc cgggacgacg tgaccctgtt catcagcgcg gtccaggacc aggtggtgcc
     3181 ggacaacacc ctggcctggg tgtgggtgcg cggcctggac gagctgtacg ccgagtggtc
     3241 ggaggtcgtg tccacgaact tccgggacgc ctccgggccg gccatgaccg agatcggcga
     3301 gcagccgtgg gggcgggagt tcgccctgcg cgacccggcc ggcaactgcg tgcacttcgt
     3361 ggccgaggag caggactgac acgtgctaaa acttcatttt taatttaaaa ggatctaggt
     3421 gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg
     3481 agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt
     3541 aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca
     3601 agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac
     3661 tgtccttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac
     3721 atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct
     3781 taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg
     3841 gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga gatacctaca
     3901 gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt
     3961 aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta
     4021 tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc
     4081 gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc
     4141 cttttgctgg ccttttgctc acatgttctt tcctgcgtta tcccctgatt ctgtggataa
     4201 ccgtattacc gcctttgagt gagctgatac cgctcgccgc agccgaacga ccgagcgcag
     4261 cgagtcagtg agcgaggaag cggaag
//
