LOCUS       sequence-255642-        3630 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Encodes a mini-transposon derived from V. cholerae Tn6677 CAST, with
            CmR cargo gene. The mutated right end introduces an MmeI recognition
            site (used for Tn-seq). Total transposon size = 977 bp..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3630)
  AUTHORS   Klompe SE, Vo PLH, Halpin-Healy TS, Sternberg SH
  TITLE     Transposon-encoded CRISPR-Cas systems direct RNA-guided DNA 
            integration.
  JOURNAL   Nature. 2019 Jul;571(7764):219-225. doi: 10.1038/s41586-019-1323-z. 
            Epub 2019 Jun 12.
  PUBMED    31189177
REFERENCE   2  (bases 1 to 3630)
  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: 
            "10.1038/s41586-019-1323-z"; journalName: "Nature"; date: "2019-07";
            volume: "571"; issue: "7764"; pages: "219-225"
COMMENT     Sequence Label: pSL0746 (pDonor_R-MmeI)
FEATURES             Location/Qualifiers
     source          1..3630
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      1..589
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     490..509
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     743..760
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    877..898
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(913..918,919..936,937..943)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    951..967
                     /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)."
     primer_bind     956..978
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     975..991
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     975..991
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             1146..1805
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(1212..1231)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     primer_bind     complement(2005..2022)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2005..2021)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2014..2036)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(2230..2249)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2349..2371
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2409..2427)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2495..2599
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2600..2668,2669..3460)
                     /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     complement(2818..2837)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
       61 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
      121 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
      181 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
      241 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
      301 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
      361 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
      421 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
      481 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
      541 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
      601 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
      661 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
      721 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcccaat
      781 acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc acgacaggtt
      841 tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc tcactcatta
      901 ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa ttgtgagcgg
      961 ataacaattt cacacaggaa acagctatga ccatgattac gccaagcttt gttggaacaa
     1021 ccataaaatg ataattacac ccataaattg ataattatca cacccataaa ttgatattgc
     1081 ctcttcatgg tctaaacttc agtaagttta cgacattttc atgaagaggt catttccggg
     1141 gatccatgga gaaaaaaatc actggatata ccaccgttga tatatcccaa tggcatcgta
     1201 aagaacattt tgaggcattt cagtcagttg ctcaatgtac ctataaccag accgttcagc
     1261 tggatattac ggccttttta aagaccgtaa agaaaaataa gcacaagttt tatccggcct
     1321 ttattcacat tcttgcccgc ctgatgaatg ctcatccgga gttccgtatg gcaatgaaag
     1381 acggtgagct ggtgatatgg gatagtgttc acccttgtta caccgttttc catgagcaaa
     1441 ctgaaacgtt ttcatcgctc tggagtgaat accacgacga tttccggcag tttctacaca
     1501 tatattcgca agatgtggcg tgttacggtg aaaacctggc ctatttccct aaagggttta
     1561 ttgagaatat gtttttcgtc tcagccaatc cctgggtgag tttcaccagt tttgatttaa
     1621 acgtggccaa tatggacaac ttcttcgccc ccgttttcac tatgggcaaa tattatacgc
     1681 aaggcgacaa ggtgctgatg ccgctggcga ttcaggttca tcatgccgtt tgtgatggct
     1741 tccatgtcgg cagaatgctt aatgaattac aacagtactg cgatgagtgg cagggcgggg
     1801 cgtaattttt ttaaggcagt tattggtgcc cttctagagt ccttactgca gtagttttgc
     1861 tgaaatactc gattcacaaa aatatcaact tatggttgtt ttgtgagata tcaatatatg
     1921 gttgttttgt ggttaagttg ctgattataa ataattatta aatatcactt tatggttgca
     1981 tcaacaggta ccgagctcga attcactggc cgtcgtttta caacgtcgtg actgggaaaa
     2041 ccctggcgtt acccaactta atcgccttgc agcacatccc cctttcgcca gctggcgtaa
     2101 tagcgaagag gcccgcaccg atcgcccttc ccaacagttg cgcagcctga atggcgaatg
     2161 gcgcctgatg cggtattttc tccttacgca tctgtgcggt atttcacacc gcatatggtg
     2221 cactctcagt acaatctgct ctgatgccgc atagttaagc cagccccgac acccgccaac
     2281 acccgctgac gcgccctgac gggcttgtct gctcccggca tccgcttaca gacaagctgt
     2341 gaccgtctcc gggagctgca tgtgtcagag gttttcaccg tcatcaccga aacgcgcgag
     2401 acgaaagggc ctcgtgatac gcctattttt ataggttaat gtcatgataa taatggtttc
     2461 ttagacgtca ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt
     2521 ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata
     2581 atattgaaaa aggaagagta tgagtattca acatttccgt gtcgccctta ttcccttttt
     2641 tgcggcattt tgccttcctg tttttgctca cccagaaacg ctggtgaaag taaaagatgc
     2701 tgaagatcag ttgggtgcac gagtgggtta catcgaactg gatctcaaca gcggtaagat
     2761 ccttgagagt tttcgccccg aagaacgttt tccaatgatg agcactttta aagttctgct
     2821 atgtggcgcg gtattatccc gtattgacgc cgggcaagag caactcggtc gccgcataca
     2881 ctattctcag aatgacttgg ttgagtactc accagtcaca gaaaagcatc ttacggatgg
     2941 catgacagta agagaattat gcagtgctgc cataaccatg agtgataaca ctgcggccaa
     3001 cttacttctg acaacgatcg gaggaccgaa ggagctaacc gcttttttgc acaacatggg
     3061 ggatcatgta actcgccttg atcgttggga accggagctg aatgaagcca taccaaacga
     3121 cgagcgtgac accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg
     3181 cgaactactt actctagctt cccggcaaca attaatagac tggatggagg cggataaagt
     3241 tgcaggacca cttctgcgct cggcccttcc ggctggctgg tttattgctg ataaatctgg
     3301 agccggtgag cgtgggtctc gcggtatcat tgcagcactg gggccagatg gtaagccctc
     3361 ccgtatcgta gttatctaca cgacggggag tcaggcaact atggatgaac gaaatagaca
     3421 gatcgctgag ataggtgcct cactgattaa gcattggtaa ctgtcagacc aagtttactc
     3481 atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct aggtgaagat
     3541 cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc
     3601 agaccccgta gaaaagatca aaggatcttc
//
