LOCUS       sequence-273330-        2559 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 2559)
  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 2559)
  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-BufAI-1
FEATURES             Location/Qualifiers
     source          1..2559
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(61..393)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITNSSSVPGDPPADICRCCRHASLALAVVLQRRDWENPGVTQ
                     LNRLAAHPPFASWRNSEEARTDRPSQQLRSLNGEWRLMRYFLLTHLCGISHRIWCTLST
                     ICSDAA"
     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(389..405)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(389..405)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(402..424)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    413..429
                     /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(437..443,444..461,462..467))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    482..503
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(620..637)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(791..1379)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(871..890)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1550..2341,2342..2410))
                     /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     2173..2192
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2411..2515)
                     /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 gcacctgcag atatctgcag gtggatcccc
      361 gggtaccgag ctcgaattcg taatcatggt catagctgtt tcctgtgtga aattgttatc
      421 cgctcacaat tccacacaac atacgagccg gaagcataaa gtgtaaagcc tggggtgcct
      481 aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa
      541 acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta
      601 ttgggcgcta ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc
      661 gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg
      721 caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt
      781 tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa
      841 gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct
      901 ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc
      961 cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg
     1021 tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct
     1081 tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag
     1141 cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga
     1201 agtggtggcc taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga
     1261 agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg
     1321 gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag
     1381 aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag
     1441 ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat
     1501 gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct
     1561 taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac
     1621 tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa
     1681 tgataccgcg agagccacgc tcaccggctc cagatttatc agcaataaac cagccagccg
     1741 gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt
     1801 gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca
     1861 ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt
     1921 cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct
     1981 tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg
     2041 cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg
     2101 agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg
     2161 cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa
     2221 aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt
     2281 aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt
     2341 gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt
     2401 gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca
     2461 tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat
     2521 ttccccgaaa agtgccacct gacgtctaag aaaccatta
//
