LOCUS       sequence-273345-        2556 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Entry vector useful for golden gate or other seamless cloning 
            technique based on Type IIS restriction enzymes.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2556)
  AUTHORS   Altaib H, Ozaki Y, Kozakai T, Badr Y, Nomura I, Suzuki T
  TITLE     A New Escherichia coli Entry Vector Series (pIIS18) for Seamless 
            Gene Cloning Using Type IIS Restriction Enzymes.
  JOURNAL   Microbiol Resour Announc. 2019 Oct 10;8(41). pii: 8/41/e00323-19. 
            doi: 10.1128/MRA.00323-19.
  PUBMED    31601653
REFERENCE   2  (bases 1 to 2556)
  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: "Microbiol 
            Resour Announc."; date: "2019-10-10"; volume: "8(41). pii: 
            8/41/e00323-19. doi"; pages: " 10.1128/MRA.00323-19"
COMMENT     Sequence Label: pIIS18-SapI
FEATURES             Location/Qualifiers
     source          1..2556
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(61..390)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITNSSSVPGDPRSSRRACRHASLALAVVLQRRDWENPGVTQL
                     NRLAAHPPFASWRNSEEARTDRPSQQLRSLNGEWRLMRYFLLTHLCGISHRIWCTLSTI
                     CSDAA"
     primer_bind     66..85
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     279..301
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     293..310
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     294..310
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(386..402)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(386..402)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(399..421)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    410..426
                     /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(434..440,441..458,459..464))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    479..500
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(617..634)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(788..1376)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(868..887)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1547..2338,2339..2407))
                     /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     2170..2189
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2408..2512)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gagcagacaa gcccgtcagg gcgcgtcagc gggtgttggc gggtgtcggg gctggcttaa
       61 ctatgcggca tcagagcaga ttgtactgag agtgcaccat atgcggtgtg aaataccgca
      121 cagatgcgta aggagaaaat accgcatcag gcgccattcg ccattcaggc tgcgcaactg
      181 ttgggaaggg cgatcggtgc gggcctcttc gctattacgc cagctggcga aagggggatg
      241 tgctgcaagg cgattaagtt gggtaacgcc agggttttcc cagtcacgac gttgtaaaac
      301 gacggccagt gccaagcttg catgcctgca ggctcttcgc gaagagcgag gatccccggg
      361 taccgagctc gaattcgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc
      421 tcacaattcc acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat
      481 gagtgagcta actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc
      541 tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg
      601 ggcgctattc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag
      661 cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag
      721 gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc
      781 tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc
      841 agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc
      901 tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt
      961 cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg
     1021 ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat
     1081 ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag
     1141 ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt
     1201 ggtggcctaa ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc
     1261 cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta
     1321 gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag
     1381 atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga
     1441 ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa
     1501 gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa
     1561 tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc
     1621 ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga
     1681 taccgcgaga gccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa
     1741 gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt
     1801 gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg
     1861 ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc
     1921 aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg
     1981 gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag
     2041 cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt
     2101 actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt
     2161 caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac
     2221 gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac
     2281 ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag
     2341 caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa
     2401 tactcatact cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga
     2461 gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc
     2521 cccgaaaagt gccacctgac gtctaagaaa ccatta
//
