LOCUS       sequence-433684-        3801 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Use together with pToxEXP/pToxAmp plasmids to insert heterologous 
            genes at ho locus.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3801)
  AUTHORS   Evans S, Lu Z, McDonnell L, Anderson W, Peralta F, Watkins T , Ahmed
            H, Luna-Flores CH, Loan T, Navone L, Trau M, Scott CH, Speight R, 
            Vickers CE, Peng B
  TITLE     The integration of tandem gene repeats via a bacterial type-II 
            toxin-antitoxin-mediated gene amplification (ToxAmp) system and 
            stability visualisation in Saccharomyces cerevisiae
  JOURNAL   bioRxiv 2024.01.30.578080
REFERENCE   2  (bases 1 to 3801)
  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: "bioRxiv 
            2024.01.30.578080"
COMMENT     Sequence Label: pITEu2
FEATURES             Location/Qualifiers
     source          1..3801
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        59..402
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     primer_bind     complement(618..634)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(618..634)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(631..653)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    642..658
                     /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(666..672,673..690,691..696))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    711..732
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(849..866)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1020..1608)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1100..1119)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1779..2570,2571..2639))
                     /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     2402..2421
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2640..2744)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     2812..2830
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(2868..2890)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     2990..3009
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3203..3225
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     3217..3234
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     3218..3234
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 ataacagggt aatatagctg tttagcttgc ctcgtccccg ccgggtcacc cggccagcga
       61 catggaggcc cagaataccc tccttgacag tcttgacgtg cgcagctcag gggcatgatg
      121 tgactgtcgc ccgtacattt agcccataca tccccatgta taatcatttg catccataca
      181 ttttgatggc cgcacggcgc gaagcaaaaa ttacggctcc tcgctgcgga cctgcgagca
      241 gggaaacgct cccctcacag acgcgttgaa ttgtccccac gccgcgcccc tgtagagaaa
      301 tataaaaggt taggatttgc cactgaggtt cttctttcat atacttcctt ttaaaatctt
      361 gctaggatac agttctcaca tcacatccga acataaacaa ccatgggtaa aaagcctgaa
      421 ctcaccgcga cgtctgtcga gaagtttctg atcgaaaagt tcgacagcgt ctccgacctg
      481 atgcagctct cggagggcga agaatctcgt gctttcagct tcgatgtagg agggcgtgga
      541 tatgtcctgc gggtaaatag ctgcgccgat ggtttctaca aagatcgtta tgtttaaacg
      601 catgcggcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc gctcacaatt
      661 ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta atgagtgagc
      721 taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc
      781 cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct
      841 tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca
      901 gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac
      961 atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt
     1021 ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg
     1081 cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc
     1141 tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc
     1201 gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc
     1261 aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac
     1321 tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt
     1381 aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct
     1441 aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc
     1501 ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt
     1561 ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg
     1621 atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc
     1681 atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa
     1741 tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag
     1801 gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg
     1861 tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga
     1921 gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag
     1981 cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa
     2041 gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc
     2101 atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca
     2161 aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg
     2221 atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat
     2281 aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc
     2341 aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg
     2401 gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg
     2461 gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt
     2521 gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca
     2581 ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata
     2641 ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac
     2701 atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa
     2761 gtgccacctg acgtctaaga aaccattatt atcatgacat taacctataa aaataggcgt
     2821 atcacgaggc cctttcgtct cgcgcgtttc ggtgatgacg gtgaaaacct ctgacacatg
     2881 cagctcccgg agacggtcac agcttgtctg taagcggatg ccgggagcag acaagcccgt
     2941 cagggcgcgt cagcgggtgt tggcgggtgt cggggctggc ttaactatgc ggcatcagag
     3001 cagattgtac tgagagtgca ccatatgcgg tgtgaaatac cgcacagatg cgtaaggaga
     3061 aaataccgca tcaggcgcca ttcgccattc aggctgcgca actgttggga agggcgatcg
     3121 gtgcgggcct cttcgctatt acgccagctg gcgaaagggg gatgtgctgc aaggcgatta
     3181 agttgggtaa cgccagggtt ttcccagtca cgacgttgta aaacgacggc cagtgcatgc
     3241 gtttaaacca cgaaaagttc accataactt cgaataaagt cgcggaaaaa agtaaacagc
     3301 tattgctact caaatgaggt ttgcagaagc ttgttgaagc atgatgaagc gttctaaacg
     3361 cactattcat cattaaatat ttaaagctca taaaattgta ttcaattcct attctaaatg
     3421 gcttttattt ctattacaac tattagctct aaatccatat cctcataagc agcaatcaat
     3481 tctatctata ctttaaaaga tctatgatta cttatcacga cgcattcgcg aaagcgaacc
     3541 attaccttga tgatgcagat ctcccggtcg tcattactct acatggacgc tttagccagg
     3601 gctggtattt ctgtttcgaa gcacgagaat ttctcgaaac tggagatgag gccgcgcgct
     3661 tagctggtaa cgcacctttt attattgata aagacagtgg tgaaattcat tctctgggaa
     3721 cggcaaaacc gctggaagaa tatctacagg attacgaaat aaaaaaggct accttcggct
     3781 tgccctgaag ttacgctagg g
//
