LOCUS       sequence-335359-        3760 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  test plasmid for Golden Gate assembly, expresses lacI repressor.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3760)
  AUTHORS   Matsumura I
  TITLE     Golden Gate assembly of BioBrick-compliant parts using Type II 
            restriction endonucleases.
  JOURNAL   Biotechniques. 2022 May;72(5):185-193. doi: 10.2144/btn-2021-0083. 
            Epub 2022 Mar 8.
  PUBMED    35255734
REFERENCE   2  (bases 1 to 3760)
  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: 
            "10.2144/btn-2021-0083"; journalName: "Biotechniques"; date: 
            "2022-05"; volume: "72"; issue: "5"; pages: "185-193"
COMMENT     Sequence Label: lacI-Ptac-lacO-BsaI-BsaI-IMBB4-pUC
FEATURES             Location/Qualifiers
     source          1..3760
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(64..83)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     protein_bind    complement(689..713)
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             complement(join(813..1604,1605..1673))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     1436..1455
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1674..1778)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     1877..1899
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     1891..1908
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     1892..1908
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_feature    1973..1994
                     /label=BioBrick prefix
                     /note="BioBrick prefix for parts that do not start with 
                     ""ATG"""
     CDS             2014..3096
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     complement(2033..2052)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(3342..3347,3348..3364,3365..3371)
                     /label=trc promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    3379..3395
                     /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)."
     RBS             3411..3419
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
     misc_feature    3455..3475
                     /label=BioBrick suffix
                     /note="universal suffix for all parts"
     primer_bind     complement(3556..3572)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3556..3572)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     rep_origin      complement(join(3744..3760,1..572))
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac
       61 tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc
      121 tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata
      181 gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc
      241 acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca
      301 acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag
      361 cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta
      421 gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg
      481 gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc
      541 agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt
      601 ctgacgctct attcaacaaa gccgccgtcc cgtcaagtca gcgtaaatgg gtagggggct
      661 tcaaatcgtc ctcgtgatac caattcggag cctgcttttt tgtacaaact tgttgataat
      721 ggcaattcaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc
      781 taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct
      841 atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata
      901 actacgatac gggagggctt accatctggc cccagtgctg caatgatacc gcgagagcca
      961 cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga
     1021 agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga
     1081 gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg
     1141 gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga
     1201 gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt
     1261 gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct
     1321 cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca
     1381 ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat
     1441 accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga
     1501 aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc
     1561 aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg
     1621 caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc
     1681 ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt
     1741 gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca
     1801 gatacctgaa acaaaaccca tcgtacggcc aaggaagtct ccaataactg tgatccacca
     1861 caagcgccag ggttttccca gtcacgacgt tgtaaaacga cggccagtca tgcataatcc
     1921 gcacgcatct ggaataagga agtgccattc cgcctgacct caattgacgc gtgaattcgc
     1981 ggccgcttct agagtactta gaggaaataa atcatgaaac cagtaacgtt atacgatgtc
     2041 gcagagtatg ccggtgtctc ttatcagacc gtttcccgcg tggtgaacca ggccagccac
     2101 gtttctgcga aaacgcggga aaaagtggaa gcggcgatgg cggagctgaa ttacattccc
     2161 aaccgcgtgg cacaacaact ggcgggcaaa cagtcgttgc tgattggcgt tgccacctcc
     2221 agtctggccc tgcacgcgcc gtcgcaaatt gtcgcggcga ttaaatctcg cgccgatcaa
     2281 ctgggtgcca gcgtggtggt gtcgatggta gaacgaagcg gcgtcgaagc ctgtaaagcg
     2341 gcggtgcaca atcttctcgc gcaacgcgtc agtgggctga tcattaacta tccgctggat
     2401 gaccaggatg ccattgctgt ggaagctgcc tgcactaatg ttccggcgtt atttcttgat
     2461 gtctctgacc agacacccat caacagtatt attttctccc atgaagacgg tacgcgactg
     2521 ggcgtggagc atctggtcgc attgggtcac cagcaaatcg cgctgttagc gggtccatta
     2581 agttctgtct cggcgcgtct gcgtctggct ggctggcata aatatctcac tcgcaatcaa
     2641 attcagccga tagcggaacg ggaaggcgac tggagtgcca tgtccggttt tcaacaaacc
     2701 atgcaaatgc tgaatgaggg catcgttccc actgcgatgc tggttgccaa cgatcagatg
     2761 gcgctgggcg caatgcgcgc cattaccgag tccgggctgc gcgttggtgc ggatatttcg
     2821 gtagtgggat acgacgatac cgaagacagc tcatgttata tcccgccgtt aaccaccatc
     2881 aaacaggatt ttcgcctgct ggggcaaacc agcgtggacc gcttgctgca actctctcag
     2941 ggccaggcgg tgaagggcaa tcagctgttg cccgtctcac tggtgaaaag aaaaaccacc
     3001 ctggcgccca atacgcaaac cgcctctccc cgcgcgttgg ccgattcatt aatgcagctg
     3061 gcacgacagg tttcccgact ggaaagcggg cagtgattga gataaaaatg ccagccgatc
     3121 gggctggcat tttgccttta gtactagagg tttgacagct tatcatcgac tgcacggtgc
     3181 accaatgctt ctggcgtcag gcagccatcg gaagctgtgg tatggctgtg caggtcgtaa
     3241 atcactgcat aattcgtgtc gctcaaggcg cactcccgtt ctggataatg ttttttgcgc
     3301 cgacatcata acggttctgg caaatattct gaaatgagct gttgacaatt aatcatccgg
     3361 ctcgtataat gtgtggaatt gtgagcggat aacaatttca cactactaga taaggaggta
     3421 tcatatggga gaccgatatc ggtctcatgg ttgatactag tagcggccgc tgcagctcga
     3481 ggtcgaccag ctcatgcata ggctaggtgg aggctcagtg atgataagtc tgcgatggtg
     3541 gatgcatgtg tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc tcagagggca
     3601 caatcctatt ccgcgctatc cgacaatctc caagacatta ggtggagttc agttcggcgt
     3661 atggcatatg tcgctggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt
     3721 aaaaaggccg cgttgctggc gtttttccat aggctccgcc
//
