LOCUS       3900_PACE_AP_GII        3659 bp    DNA     circular SYN 31-MAY-2023
DEFINITION  .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  recombinant plasmid
REFERENCE   1  (bases 1 to 3659)
  AUTHORS   Nicholas Southern
  TITLE     Direct Submission
  JOURNAL   Exported Jun 24, 2025 from SnapGene 8.0.3
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..3659
                     /mol_type="other DNA"
                     /organism="recombinant plasmid"
     modified_base   complement(1..16)
                     /label=USER junction
                     /note="USER junction"
                     /note="color: #a6acb3; direction: LEFT"
     CDS             complement(17..676)
                     /label=cat (CmR)
                     /note="cat (CmR)"
                     /note="from pACYCDuet-1"
                     /note="color: #993366; direction: LEFT"
     modified_base   92
                     /label=mutation
                     /note="mutation"
                     /note="annotated as a G in pACYCDuet cat markerannotation, 
                     here it is an A, but this mutation is silent from codon GTC
                     (Val, 25% codon usage) to GTT (Val, 21% usage), so it 
                     should not be of functional relevance"
                     /note="color: #a6acb3"
     promoter        677..770
                     /note="cat promoter"
                     /note="from pACYCDuet-1"
                     /note="color: #ffffff; direction: RIGHT"
     misc_feature    complement(775..791)
                     /label=USER linker
                     /note="USER linker"
                     /note="color: #a6acb3; direction: LEFT"
     terminator      792..1216
                     /note="rrnB1 transcriptional terminator"
                     /note="color: #ffffff"
     misc_feature    complement(1224..1236)
                     /label=USER linker
                     /note="USER linker"
                     /note="color: #a6acb3; direction: LEFT"
     enhancer        1237..1284
                     /note="Protected by pspF"
                     /note="color: #a6acb3"
     protein_bind    1237..1251
                     /note="UAS II"
                     /note="color: #31849b"
     protein_bind    1239..1248
                     /label=pspF
                     /note="/%_Identity=100"
                     /note="/Motif=TGAAATTCGC"
                     /note="/annotation_group=TGAAATTCGC: 6,220 -> 6,229"
                     /note="/modified_by=tinawang"
                     /note="color: #31849b"
     RBS             1244..1253
                     /label=pspF RBS
                     /note="pspF RBS"
                     /note="color: #a6acb3"
     prim_transcript 1253
                     /note="color: #a6acb3; direction: RIGHT"
     protein_bind    1255..1273
                     /note="UAS I"
                     /note="color: #31849b"
     promoter        complement(1257..1284)
                     /note="pspF P2"
                     /note="color: #ffffff; direction: LEFT"
     -10_signal      complement(1257..1262)
                     /label=-10
                     /note="-10"
                     /note="color: #a6acb3; direction: LEFT"
     protein_bind    1259..1268
                     /label=pspF
                     
                     /note="/%_Identity=100"
                     /note="/Motif=TGTAATTCGC"
                     /note="/annotation_group=TGTAATTCGC: 6,240 -> 6,249"
                     /note="/modified_by=tinawang"
                     /note="color: #31849b"
     prim_transcript 1269
                     /note="color: #a6acb3; direction: RIGHT"
     promoter        complement(1275..1303)
                     /note="pspF P1"
                     /note="color: #ffffff; direction: LEFT"
     -10_signal      complement(1275..1280)
                     /label=-10
                     /note="-10"
                     /note="color: #a6acb3; direction: LEFT"
     -35_signal      complement(1279..1284)
                     /label=-35
                     /note="-35"
                     /note="color: #a6acb3; direction: LEFT"
     -35_signal      complement(1298..1303)
                     /label=-35
                     /note="-35"
                     /note="color: #a6acb3; direction: LEFT"
     protein_bind    1301..1333
                     /note="High Affinity IHF Site"
                     /note="color: #31849b"
     promoter        1335..1351
                     /note="Sigma54 Core Promoter"
                     /note="color: #ffffff"
     -35_signal      1335..1342
                     /label=-35
                     /note="/created_by=tinawang"
                     /note="color: #a6acb3"
     -35_signal      1337..1338
                     /label=-24
                     /note="-24"
                     /note="color: #a6acb3"
     -10_signal      1347..1351
                     /label=-10
                     /note="/created_by=tinawang"
                     /note="color: #a6acb3"
     -10_signal      1349..1350
                     /label=-12
                     /note="-12"
                     /note="color: #a6acb3"
     protein_bind    1363..1382
                     /label=lac operator (symmetric)
                     /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). The 
                     symmetric lac operator was optimized for tight binding of 
                     lac repressor."
                     /note="color: #31849b"
     prim_transcript 1384
                     /note="color: #a6acb3; direction: RIGHT"
     promoter        complement(1391..1418)
                     /note="pspF P3"
                     /note="color: #ffffff; direction: LEFT"
     -10_signal      complement(1391..1396)
                     /label=-10
                     /note="-10"
                     /note="color: #a6acb3; direction: LEFT"
     -35_signal      complement(1413..1418)
                     /label=-35
                     /note="-35"
                     /note="color: #a6acb3; direction: LEFT"
     modified_base   1423..1437
                     /label=USER junction
                     /note="USER junction"
                     /note="color: #a6acb3"
     RBS             1438..1451
                     /note="sd8 RBS (from Ringquist and Gold  Mol. Micro.1992)"
                     /note="color: #a6acb3"
     CDS             1452..2726
                     /codon_start=1
                     /label=III
                     /note="/dnas_title=III"
                     /note="/vntifkey=4"
                     /note="color: #993366"
                     /translation="MKKLLFAIPLVVPFYSHSAETVESCLAKPHTENSFTNVWKDDKTL
                     DRYANYEGCLWNATGVVVCTGDETQCYGTWVPIGLAIPENEGGGSEGGGSEGGGSEGGG
                     TKPPEYGDTPIPGYTYINPLDGTYPPGTEQNPANPNPSLEESQPLNTFMFQNNRFRNRQ
                     GALTVYTGTVTQGTDPVKTYYQYTPVSSKAMYDAYWNGKFRDCAFHSGFNEDPFVCEYQ
                     GQSSDLPQPPVNAGGGSGGGSGGGSEGGGSEGGGSEGGGSEGGGSGGGSGSGDFDYEKM
                     ANANKGAMTENADENALQSDAKGKLDSVATDYGAAIDGFIGDVSGLANGNGATGDFAGS
                     NSQMAQVGDGDNSPLMNNFRQYLPSLPQSVECRPFVFGAGKPYEFSIDCDKINLFRGVF
                     AFLLYVATFMYVFSTFANILRNKES"
     misc_feature    2594..2639
                     /label=modified to remove internal promoter
                     /note="modified to remove internal promoter"
                     /note="color: #a6acb3"
     modified_base   complement(2727..2741)
                     /label=USER junction
                     /note="USER junction"
                     /note="color: #a6acb3; direction: LEFT"
     misc_difference complement(2742..2748)
                     /label=cloning scar
                     /note="cloning scar"
                     /note="color: #a6acb3; direction: LEFT"
     terminator      complement(2749..2793)
                     /note="rrnB1 transcriptional terminator"
                     /note="color: #ffffff; direction: LEFT"
     modified_base   complement(2794..2808)
                     /label=USER junction
                     /note="USER junction"
                     /note="color: #a6acb3; direction: LEFT"
     modified_base   complement(2854..2871)
                     /label=User Junction
                     /note="User Junction"
                     /note="color: #a6acb3; direction: LEFT"
     rep_origin      2883..3621
                     /label=cloDF13
                     /note="/dnas_title=cloDF13"
                     /note="/vntifkey=33"
                     /note="color: #ffff00"
ORIGIN
        1 acggcctatt tggcttttac gccccgccct gccactcatc gcagtactgt tgtaattcat
       61 taagcattct gccgacatgg aagccatcac aaacggcatg atgaacctga atcgccagcg
      121 gcatcagcac cttgtcgcct tgcgtataat atttgcccat agtgaaaacg ggggcgaaga
      181 agttgtccat attggccacg tttaaatcaa aactggtgaa actcacccag ggattggctg
      241 agacgaaaaa catattctca ataaaccctt tagggaaata ggccaggttt tcaccgtaac
      301 acgccacatc ttgcgaatat atgtgtagaa actgccggaa atcgtcgtgg tattcactcc
      361 agagcgatga aaacgtttca gtttgctcat ggaaaacggt gtaacaaggg tgaacactat
      421 cccatatcac cagctcaccg tctttcattg ccatacggaa ctccggatga gcattcatca
      481 ggcgggcaag aatgtgaata aaggccggat aaaacttgtg cttatttttc tttacggtct
      541 ttaaaaaggc cgtaatatcc agctgaacgg tctggttata ggtacattta gcaactgact
      601 gaaatgcctc aaaatgttct ttacgatgcc attgggatat atcaacggtg gtatatccag
      661 tgattttttt ctccatttta gcttccttag ctcctgaaaa tctcgataac tcaaaaaata
      721 cgcccggtag tgatcttatt tcattatggt gaaagttgga acctcttacg tgccacagtc
      781 agtagggccc tggctgtttt ggcggatgag agaagatttt cagcctgata cagattaaat
      841 cagaacgcag aagcggtctg ataaaacaga atttgcctgg cggcagtagc gcggtggtcc
      901 cacctgaccc catgccgaac tcagaagtga aacgccgtag cgccgatggt agtgtggggt
      961 ctccccatgc gagagtaggg aactgccagg catcaaataa aacgaaaggc tcagtcgaaa
     1021 gactgggcct ttcgttttat ctgttgtttg tcggtgaacg ctctcctgag taggacaaat
     1081 ccgccgggag cggatttgaa cgttgcgaag caacggcccg gagggtggcg ggcaggacgc
     1141 ccgccataaa ctgccaggca tcaaattaag cagaaggcca tcctgacgga tggccttttt
     1201 gcgtttctac aaactcgctg aggacattgg ctccgtgatg aaattcgcca cttgttagtg
     1261 taattcgcta actcatcctg gcatgttgct gttgattctt caatcagatc tttataaatc
     1321 aaaaagataa aaaattggca cgcaaattgt attaacagtt caaattgtga gcgctcacaa
     1381 ttgcaggaca atcctgaacg cagaaatcaa gaggacaaca ttacctgcag gtgcagtaaa
     1441 ggaaaaaaaa aatgaaaaaa ttattattcg caattccttt agttgttcct ttctattctc
     1501 actccgctga aactgttgaa agttgtttag caaaacccca tacagaaaat tcatttacta
     1561 acgtctggaa agacgacaaa actttagatc gttacgctaa ctatgagggc tgtctgtgga
     1621 atgctacagg cgttgtagtt tgtactggtg acgaaactca gtgttacggt acatgggttc
     1681 ctattgggct tgctatccct gaaaatgagg gtggtggctc tgagggtggc ggttctgagg
     1741 gtggcggttc tgagggtggc ggtactaaac ctcctgagta cggtgataca cctattccgg
     1801 gctatactta tatcaaccct ctcgacggca cttatccgcc tggtactgag caaaaccccg
     1861 ctaatcctaa tccttctctt gaggagtctc agcctcttaa tactttcatg tttcagaata
     1921 ataggttccg aaataggcag ggggcattaa ctgtttatac gggcactgtt actcaaggca
     1981 ctgaccccgt taaaacttat taccagtaca ctcctgtatc atcaaaagcc atgtatgacg
     2041 cttactggaa cggtaaattc agagactgcg ctttccattc tggctttaat gaggatccat
     2101 tcgtttgtga atatcaaggc caatcgtctg acctgcctca acctcctgtc aatgctggcg
     2161 gcggctctgg tggtggttct ggtggcggct ctgagggtgg tggctctgag ggtggcggtt
     2221 ctgagggtgg cggctctgag ggaggcggtt ccggtggtgg ctctggttcc ggtgattttg
     2281 attatgaaaa gatggcaaac gctaataagg gggctatgac cgaaaatgcc gatgaaaacg
     2341 cgctacagtc tgacgctaaa ggcaaacttg attctgtcgc tactgattac ggtgctgcta
     2401 tcgatggttt cattggtgac gtttccggcc ttgctaatgg taatggtgct actggtgatt
     2461 ttgctggctc taattcccaa atggctcaag tcggtgacgg tgataattca cctttaatga
     2521 ataatttccg tcaatattta ccttccctcc ctcaatcggt tgaatgtcgc ccttttgtct
     2581 ttggcgctgg taaaccttac gagttcagta tcgactgcga taagatcaac ctgttccgcg
     2641 gtgtctttgc gtttctttta tatgttgcca cctttatgta tgtattttct acgtttgcta
     2701 acatactgcg taataaggag tcttaaaggg tccctaagtc tcctcagcaa aacgaaaggc
     2761 ccagtctttc gactgagcct ttcgttttat ttgaccggat gtcctcttcc tcctgggctg
     2821 acttcaggtg ctacatttga agagataaat tgcactgaaa tctagagcgg ttcagtagaa
     2881 aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatcttttg ccctgtaaac
     2941 gaaaaaacca cctggggagg tggtttgatc gaaggttaag tcagttgggg aactgcttaa
     3001 ccgtggtaac tggctttcgc agagcacagc aaccaaatct gtccttccag tgtagccgga
     3061 ctttggcgca cacttcaaga gcaaccgcgt gtttagctaa acaaatcctc tgcgaactcc
     3121 cagttaccaa tggctgctgc cagtggcgtt ttaccgtgct tttccgggtt ggactcaagt
     3181 gaacagttac cggataaggc gcagcagtcg ggctgaacgg ggagttcttg cttacagccc
     3241 agcttggagc gaacgaccta caccgagccg agataccagt gtgtgagcta tgagaaagcg
     3301 ccacacttcc cgtaagggag aaaggcggaa caggtatccg gtaaacggca gggtcggaac
     3361 aggagagcgc aagagggagc gacccgccgg aaacggtggg gatctttaag tcctgtcggg
     3421 tttcgcccgt actgtcagat tcatggttga gcctcacggc tcccacagat gcaccggaaa
     3481 agcgtctgtt tatgtgaact ctggcaggag ggcggagcct atggaaaaac gccaccggcg
     3541 cggccctgct gttttgcctc acatgttagt cccctgctta tccacggaat ctgtgggtaa
     3601 ctttgtatgt gtccgcagcg cccgccgcag tctcacgccc ggagcgtagc gaccgagtg
//
