LOCUS       sequence-72935-e        4598 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Golden Gate entry vector at position 2, LacZ, final entry vector for
            the assembly.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4598)
  AUTHORS   Kirchmaier S, Lust K, Wittbrodt J
  TITLE     Golden GATEway cloning--a combinatorial approach to generate fusion 
            and recombination constructs.
  JOURNAL   PLoS One. 2013 Oct 7;8(10):e76117. doi: 
            10.1371/journal.pone.0076117.
  PUBMED    24116091
REFERENCE   2  (bases 1 to 4598)
  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: "PLoS One.";
            date: "2013-10-7"; volume: "8(10):e76117. doi"; pages: " 
            10.1371/journal.pone.0076117"
COMMENT     Sequence Label: pGGEV_2'_XcmI-LacZ
FEATURES             Location/Qualifiers
     source          1..4598
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(1..456)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(138..159)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     350..369
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        971..989
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     CDS             complement(1027..1410)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITPSYLGDTIEYSSYASNALGALPYGRPAGGREFTSDIEFPR
                     PPWRPGACDVGPNSPYSESYYNSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEA
                     RTDRPSQQLRSLNGEWTRPVAAH"
     primer_bind     1176..1198
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     1190..1207
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     1191..1207
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1214..1233
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1214..1232
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      1263..1272
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      1267..1276
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(1370..1388)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(1406..1422)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1406..1422)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1419..1441)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1430..1446
                     /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(join(1454..1460,1461..1478,1479..1484))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1499..1520
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     intron          1746..2318
                     /label=beta-globin intron
                     /note="intron from rabbit beta-globin gene"
     polyA_signal    2391..2525
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2415..2434)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2469..2488
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     rep_origin      complement(2848..3436)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(join(3607..4398,4399..4467))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAISMSDNTAANLLLTTIGGPKELTAFFHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     4230..4249
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4468..4572)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 aaattgtaaa cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat
       61 tttttaacca ataggccgaa atcggcaaaa tcccttataa atcaaaagaa tagaccgaga
      121 tagggttgag tgttgttcca gtttggaaca agagtccact attaaagaac gtggactcca
      181 acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa ccatcaccct
      241 aatcaagttt tttggggtcg aggtgccgta aagcactaaa tcggaaccct aaagggagcc
      301 cccgatttag agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa gggaagaaag
      361 cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc gtaaccacca
      421 cacccgccgc gcttaatgcg ccgctacagg gcgcgtccca ttcgccattc aggctacgca
      481 actgttggga agggcgatcg gtgcgggcct cttcgctatt acgccagggc gcctccccca
      541 ttgacgtcaa ttacggtaaa tggcccgcct ggctcaatgc ccattgacgt caataggacc
      601 acccaccatt gacgtcaatg ggatggctca ttgcccattc atatccgttc tcacgccccc
      661 tattgacgtc aatgacggta aatggcccac ttggcagtac atcaatatct attaatagta
      721 acttggcaag tacattacta ttggaagtac gccagggtac attggcagta ctcccattga
      781 cgtcaatggc ggtaaatggc ccgcgatggc tgccaagtac atccccattg acgtcaatgg
      841 ggaggggcaa tgacgcaaat gggcgttcca ttgacgtaaa tgggcggtag gcgtgcctaa
      901 tgggaggtct atataagcaa tgctcgttta gggaaccgcc attctgcctg gggacgtcgg
      961 agcaagcttg atttaggtga cactatagag gtctcaggct ccggatccag cggtggcgtg
     1021 gcgcgcttaa tgcgccgcta cagggcgcgt ccattcgcca ttcaggctgc gcaactgttg
     1081 ggaagggcga tcggtgcggg cctcttcgct attacgccag ctggcgaaag ggggatgtgc
     1141 tgcaaggcga ttaagttggg taacgccagg gttttcccag tcacgacgtt gtaaaacgac
     1201 ggccagtgaa ttgtaatacg actcactata gggcgaattg ggcccgacgt cgcatgctcc
     1261 cggccgccat ggcggccgcg ggaattcgat atcactagtg aattcgcggc cgcctgcagg
     1321 tcgaccatat gggagagctc ccaacgcgtt ggatgcatag cttgagtatt ctatagtgtc
     1381 acctaaatag cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc
     1441 tcacaattcc acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat
     1501 gagtgagcta actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc
     1561 tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg
     1621 ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc gccacgccac cgctggtacc
     1681 taagtgagac cctcatcctc ccgggtaact aagtaaggat ctcgaccgat cctgagaact
     1741 tcagggtgag tttggggacc cttgattgtt ctttcttttt cgctattgta aaattcatgt
     1801 tatatggagg gggcaaagtt ttcagggtgt tgtttagaat gggaagatgt cccttgtatc
     1861 accatggacc ctcatgataa ttttgtttct ttcactttct actctgttga caaccattgt
     1921 ctcctcttat tttcttttca ttttctgtaa ctttttcgtt aaactttagc ttgcatttgt
     1981 aacgaatttt taaattcact tttgtttatt tgtcagattg taagtacttt ctctaatcac
     2041 ttttttttca aggcaatcag ggtatattat attgtacttc agcacagttt tagagaacaa
     2101 ttgttataat taaatgataa ggtagaatat ttctgcatat aaattctggc tggcgtggaa
     2161 atattcttat tggtagaaac aactacaccc tggtcatcat cctgcctttc tctttatggt
     2221 tacaatgata tacactgttt gagatgagga taaaatactc tgagtccaaa ccgggcccct
     2281 ctgctaacca tgttcatgcc ttcttctctt tcctacagct cctgggcaac gtgctggttg
     2341 ttgtgctgtc tcatcatttt ggcaaagaat taattcctcg acggatcgta gatccagaca
     2401 tgataagata cattgatgag tttggacaaa ccacaactag aatgcagtga aaaaaatgct
     2461 ttatttgtga aatttgtgat gctattgctt tatttgtaac cattataagc tgcaataaac
     2521 aagttaacaa caacaattgc attcatttta tgtttcaggt tcagggggag gtgtgggagg
     2581 ttttttaatt cgcggccgct ctagaactag tctgcattaa tgaatcggcc aacgcgcggg
     2641 gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc
     2701 ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac
     2761 agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa
     2821 ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc ggcccccctg acgagcatca
     2881 caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc
     2941 gttcccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata
     3001 cctgtccgcc tttctccctt cgggaagcgt ggcgctttct caatgctcac gctgtaggta
     3061 tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca
     3121 gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga
     3181 cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg
     3241 tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagga cagtatttgg
     3301 tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg
     3361 caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag
     3421 aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa
     3481 cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat
     3541 ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc
     3601 tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc
     3661 atccatagtt gcctgactgc ccgtcgtgta gataactacg atacgggagg gcttaccatc
     3721 tggccccagt gctgcaatga taccgcggga cccacgctca ccggctccag atttatcagc
     3781 aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc
     3841 catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt
     3901 gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc
     3961 ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgaaa
     4021 aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt
     4081 atcactcatg cttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg
     4141 cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc
     4201 gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa
     4261 agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt
     4321 gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt
     4381 caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag
     4441 ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta
     4501 tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat
     4561 aggggttccg cgcacatttc cccgaaaagt gccacctg
//
