LOCUS       sequence-482516-        3479 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3479)
  TITLE     Mini-Golden Subcloning Platform
REFERENCE   2  (bases 1 to 3479)
  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"
COMMENT     Sequence Label: BsaI-MCS-cloning vector for golden gate
FEATURES             Location/Qualifiers
     source          1..3479
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(329..1120,1121..1189))
                     /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     952..971
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1190..1294)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(1320..1775)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1457..1478)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1669..1688
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1902..1924
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     1916..1933
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     1917..1933
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1943..1962
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1943..1961
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     misc_feature    join(1977..1988,1989..2151)
                     /label=Tol2 right end
                     /note="right end of the Tol2 transposon from the medaka 
                     fish Oryzias latipes (Kawakami, 2007)"
     misc_feature    join(2322..2509,2510..2521)
                     /label=Tol2 left end
                     /note="left end of the Tol2 transposon from the medaka fish
                     Oryzias latipes (Kawakami, 2007)"
     primer_bind     complement(2555..2571)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2555..2571)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     primer_bind     complement(2608..2628)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(2608..2626)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2647..2663)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2647..2663)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2660..2682)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2671..2687
                     /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(2695..2701,2702..2719,2720..2725))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2740..2761
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2878..2895)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(join(3049..3479,1..158))
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3129..3148)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa
       61 gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt
      121 gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta
      181 cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat
      241 caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa
      301 gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct
      361 cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta
      421 cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcgg gacccacgct
      481 caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg
      541 gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa
      601 gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt
      661 cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta
      721 catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca
      781 gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta
      841 ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct
      901 gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg
      961 cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac
     1021 tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact
     1081 gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa
     1141 atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt
     1201 ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat
     1261 gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa atgccaccta
     1321 aattgtaagc gttaatattt tgttaaaatt cgcgttaaat ttttgttaaa tcagctcatt
     1381 ttttaaccaa taggccgaaa tcggcaaaat cccttataaa tcaaaagaat agaccgagat
     1441 agggttgagt gttgttccag tttggaacaa gagtccacta ttaaagaacg tggactccaa
     1501 cgtcaaaggg cgaaaaaccg tctatcaggg cgatggccca ctacgtgaac catcacccta
     1561 atcaagtttt ttggggtcga ggtgccgtaa agcactaaat cggaacccta aagggagccc
     1621 ccgatttaga gcttgacggg gaaagccggc gaacgtggcg agaaaggaag ggaagaaagc
     1681 gaaaggagcg ggcgctaggg cgctggcaag tgtagcggtc acgctgcgcg taaccaccac
     1741 acccgccgcg cttaatgcgc cgctacaggg cgcgtcccat tcgccattca ggctgcgcaa
     1801 ctgttgggaa gggcgatcgg tgcgggcctc ttcgctatta cgccagctgg cgaaaggggg
     1861 atgtgctgca aggcgattaa gttgggtaac gccagggttt tcccagtcac gacgttgtaa
     1921 aacgacggcc agtgagcgcg cgtaatacga ctcactatag ggcgaattgg gtaccacaga
     1981 ggtgtaaaaa gtactcaaaa attttactca agtgaaagta caagtactta gggaaaattt
     2041 tactcaatta aaagtaaaag tatctggcta gaatcttact tgagtaaaag taaaaaagta
     2101 ctccattaaa attgtacttg agtattaagg aagtaaaagt aaaagcaaga aagaaaacta
     2161 gagattcttg aagctttagg gataacaggg taatctcgag cagtagagac cgaattcatc
     2221 gatggtctct ccaaggatcc tgcagatctg tacatatgca tgggcccgta gggataacag
     2281 ggtaatcaat tggtttggta atagcaaggg aaaatagaat gaagtgatct ccaaaaaata
     2341 agtacttttt gactgtaaat aaaattgtaa ggagtaaaaa gtactttttt ttctaaaaaa
     2401 atgtaattaa gtaaaagtaa aagtattgat ttttaattgt actcaagtaa agtaaaaatc
     2461 cccaaaaata atacttaagt acagtaatca agtaaaatta ctcaagtact ttacacctct
     2521 ggttcttgac cccctacctt cagcaagccc agcagatcca ctagttctag agcggccgcc
     2581 accgcggtgg agctccagct tttgttccct ttagtgaggg ttaattgcgc gcttggcgta
     2641 atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat
     2701 acgagccgga agcataaagt gtaaagcctg gggtgcctaa tgagtgagct aactcacatt
     2761 aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta
     2821 atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc
     2881 gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa
     2941 ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa
     3001 aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct
     3061 ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac
     3121 aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc
     3181 gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc
     3241 tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg
     3301 tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga
     3361 gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta acaggattag
     3421 cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggct
//
