LOCUS       PB_CA_retrofitte        3863 bp    DNA        circular     17-JAN-2024
COMMENT     ApEinfo:methylated:1
FEATURES             Location/Qualifiers
     rep_origin      1..456
                     /direction=RIGHT
                     /note="f1 bacteriophage originofreplication;arrowindicates
                     direction of (+) strand synthesis"
                     /locus_tag="f1 ori"
     primer_bind     complement(138..159)
                     /note="F1 origin, forward primer"
                     /locus_tag="F1ori-F"
     primer_bind     350..369
                     /note="F1 origin, reverse primer"
                     /locus_tag="F1ori-R"
     primer_bind     583..605
                     /note="In lacZ gene"
                     /locus_tag="M13/pUC Forward"
     primer_bind     597..614
                     /note="In lacZ gene. Also called M13-F20orM13(-21)Forward"
                     /locus_tag="M13 Forward"
     primer_bind     598..614
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /locus_tag="M13 fwd"
     promoter        621..639
                     /note="promoter for bacteriophage T7 RNA polymerase"
                     /locus_tag="T7 promoter"
     misc_feature    complement(653..1103)
                     /note="3' piggyBac sequence"
                     /locus_tag="3' PB sequence"
     misc_feature    702..705
                     /note="TTAA"
                     /locus_tag="TTAA"
     misc_feature    706..718
                     /note="13 bp terminal inverted repeat"
                     /locus_tag="13-bp TIR"
     misc_feature    719..749
                     /note="31 bp spacer"
                     /locus_tag="31-bp spacer"
     misc_feature    750..768
                     /note="19 bp internal inverted repeat"
                     /locus_tag="19-bp IR"
     misc_feature    complement(1129..1656)
                     /note="5' piggyBac sequence"
                     /locus_tag="5' PB sequence"
     misc_feature    1490..1508
                     /note="19 bp internal inverted repeat"
                     /locus_tag="19-bp IR(1)"
     misc_feature    1509..1511
                     /note="3 bp spacer"
                     /locus_tag="3-bp spacer"
     misc_feature    1512..1524
                     /note="13 bp terminal inverted repeat"
                     /locus_tag="13-bp TIR(1)"
     misc_feature    1525..1528
                     /note="TTAA"
                     /locus_tag="TTAA(1)"
     promoter        complement(1673..1691)
                     /note="promoter for bacteriophage T3 RNA polymerase"
                     /locus_tag="T3 promoter"
     primer_bind     complement(1712..1728)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /locus_tag="M13 rev"
     primer_bind     complement(1712..1728)
                     /note="In lacZ gene. Also called M13-rev"
                     /locus_tag="M13 Reverse"
     primer_bind     complement(1725..1747)
                     /note="In lacZ gene"
                     /locus_tag="M13/pUC Reverse"
     protein_bind    complement(1736..1752)
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lacoperatortoinhibit
                     transcription in E. coli. This inhibition can be relieved
                     by adding lactose or
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
                     /locus_tag="lac operator"
     promoter        complement(1760..1790)
                     /note="promoter for the E. coli lac operon"
                     /locus_tag="lac promoter"
     protein_bind    complement(1805..1826)
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription inthepresenceof
                     cAMP."
                     /locus_tag="CAP binding site"
     primer_bind     complement(1943..1960)
                     /note="L4440 vector, forward primer"
                     /locus_tag="L4440"
     rep_origin      2114..2702
                     /direction=RIGHT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /locus_tag="ori"
     primer_bind     complement(2194..2213)
                     /note="pBR322 origin, forward primer"
                     /locus_tag="pBR322ori-F"
     CDS             complement(2873..3733)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /note="confers resistance to ampicillin, carbenicillin,and
                     related antibiotics"
                     /translation=MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEY
                     SPVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLD
                     RWEPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLL
                     RSALPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAE
                     IGASLIKHW
                     /locus_tag="AmpR"
     primer_bind     3496..3515
                     /note="Ampicillin resistance gene, reverse primer"
                     /locus_tag="Amp-R"
     promoter        complement(3734..3838)
                     /gene="bla"
                     /locus_tag="AmpR promoter"
     misc_feature    1028..1028
                     /note="A to G mutation"
                     /locus_tag="A to G mutation"
ORIGIN
        1 aaattgtaag cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat
       61 tttttaacca ataggccgaa atcggcaaaa tcccttataa atcaaaagaa tagaccgaga
      121 tagggttgag tgttgttcca gtttggaaca agagtccact attaaagaac gtggactcca
      181 acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa ccatcaccct
      241 aatcaagttt tttggggtcg aggtgccgta aagcactaaa tcggaaccct aaagggagcc
      301 cccgatttag agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa gggaagaaag
      361 cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc gtaaccacca
      421 cacccgccgc gcttaatgcg ccgctacagg gcgcgtccca ttcgccattc aggctgcgca
      481 actgttggga agggcgatcg gtgcgggcct cttcgctatt acgccagctg gcgaaagggg
      541 gatgtgctgc aaggcgatta agttgggtaa cgccagggtt ttcccagtca cgacgttgta
      601 aaacgacggc cagtgaattg taatacgact cactataggg cgaattggag ctcggtattc
      661 acgacagcag gctgaataat aaaaaaatta gaaactatta tttaacccta gaaagataat
      721 catattgtga cgtacgttaa agataatcat gcgtaaaatt gacgcatgtg ttttatcggt
      781 ctgtatatcg aggtttattt attaatttga atagatatta agttttatta tatttacact
      841 tacatactaa taataaattc aacaaacaat ttatttatgt ttatttattt attaaaaaaa
      901 aacaaaaact caaaatttct tctataaagt aacaaaactt ttaaacattc tctcttttac
      961 aaaaataaac ttattttgta ctttaaaaac agtcatgttg tattataaaa taagtaatta
     1021 gcttaaccta tacataatag aaacaaatta tacttattag tcagtcagaa acaactttgg
     1081 cacatatcaa tattatgctc tcgttaatac cggtcagtaa gcttctagcc aacaagctcg
     1141 tcatcgcttt gcagaagagc agagaggata tgctcatcgt ctaaagaact acccatttta
     1201 ttatatatta gtcacgatat ctataacaag aaaatatata tataataagt tatcacgtaa
     1261 gtagaacatg aaataacaat ataattatcg tatgagttaa atcttaaaag tcacgtaaaa
     1321 gataatcatg cgtcattttg actcacgcgg tcgttatagt tcaaaatcag tgacacttac
     1381 cgcattgaca agcacgcctc acgggagctc caagcggcga ctgagatgtc ctaaatgcac
     1441 agcgacggat tcgcgctatt tagaaagaga gagcaatatt tcaagaatgc atgcgtcaat
     1501 tttacgcaga ctatctttct agggttaaaa aagatttgcg ctttactcga cctaaacttt
     1561 aaacacgtca tagaatcttc gtttgacaaa aaccacattg tggccaagct gtgtgacgcg
     1621 acgcgcgcta aagaatggca aaccaagtcg cgcgaggtac cagcttttgt tccctttagt
     1681 gagggttaat ttcgagcttg gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt
     1741 atccgctcac aattccacac aacatacgag ccggaagcat aaagtgtaaa gcctggggtg
     1801 cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct ttccagtcgg
     1861 gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc
     1921 gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc
     1981 ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata
     2041 acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg
     2101 cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct
     2161 caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa
     2221 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc
     2281 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt
     2341 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg
     2401 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg
     2461 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct
     2521 tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc
     2581 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg
     2641 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc
     2701 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt
     2761 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa
     2821 aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat
     2881 gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct
     2941 gactccccgt cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg
     3001 caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag
     3061 ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta
     3121 attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg
     3181 ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg
     3241 gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct
     3301 ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta
     3361 tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg
     3421 gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc
     3481 cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg
     3541 gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga
     3601 tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg
     3661 ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat
     3721 gttgaatact catactcttc ctttttcaat attattgaag catttatcag ggttattgtc
     3781 tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca
     3841 catttccccg aaaagtgcca cct   
//
