LOCUS       sequence-266676-        7382 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  An integration vector for the chromosomal deletion of the upp gene 
            in P. putida KT2440; carries the upp deletion cassette..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7382)
  AUTHORS   Graf N, Altenbuchner J
  TITLE     Development of a method for markerless gene deletion in Pseudomonas 
            putida.
  JOURNAL   Appl Environ Microbiol. 2011 Aug;77(15):5549-52. doi: 
            10.1128/AEM.05055-11. Epub 2011 Jun 10.
  PUBMED    21666018
REFERENCE   2  (bases 1 to 7382)
  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: 
            "10.1128/AEM.05055-11"; journalName: "Appl Environ Microbiol"; date:
            "2011-08"; volume: "77"; issue: "15"; pages: "5549-52"
COMMENT     Sequence Label: pJOE6227.1
FEATURES             Location/Qualifiers
     source          1..7382
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(3926..4720)
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4037..4056)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     4647..4666
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     complement(5089..5105)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5089..5105)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5102..5124)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5113..5129
                     /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(5137..5143,5144..5161,5162..5167))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5182..5203
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5320..5337)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5491..6079)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5571..5590)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6250..7041,7042..7110))
                     /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     6873..6892
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7111..7215)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7356..7374
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 ctcgcgcgtt tcggtgatga cggtgaaaac ctctgacaca tgcagctccc ggagacggtc
       61 acagcttgtc tgtaagcgga tgccgggagc agacaagctg tgaccgtctc cgggagctgc
      121 atgtgtcaga ggttttcacc gtcatcaccg aaacgcgcga ggcagaacag tcggtacggc
      181 tgaccatcgg gtgccagtgc gggagtttcg ttcagcactg tcctgctcct tgtgatggtt
      241 tacaaacgta aaaagtctct ttaatacctg tttttgcttc atattgttca gcgacagctt
      301 gctgtacggc aggcaccagc tcttccggga tcagcgcgac gatacagccg ccaaatccgc
      361 cgccggtcat gcgtacgcca cctttgtcgc caatcacagc tttgacgatt tctaccagag
      421 tgtcaatttg cggcacggtg atttcgaaat catcgcgcat agaggcatga gactccgcca
      481 tcaactcgcc catacgtttc aggtcgcctt gctccagcgc gctggcagct tcaacggtgc
      541 gggcgttttc agtcagtata tgacgcacgc gttttgccac gatcgggtcc agttcatgcg
      601 caacagcgtt gaactcttca atggtgacat cacgcagggc tggctgctgg aagaaacgcg
      661 caccggtttc gcactgttca cgacgggtgt tgtattcgct gccaaccagg gtacgtttga
      721 agttactgtt gatgatgacg acagccacac ctttgggcat ggaaactgct ttggtcccca
      781 gtgagcggca atcgatcagc aaggcatgat ctttcttgcc gagcgcggaa attagctgat
      841 ccatgatccc gcagttacag cctacaaact ggttttctgc ttcctgaccg ttaagcgcga
      901 tttgtgcgcc gtccagcggc agatgataaa gctgctgcaa tacggttccg accgcgactt
      961 ccagtgaagc ggaagaactt aacccggcac cctgcggcac attgccgctg atcaccatgt
     1021 ccacgccgcc gaagctgttg ttacgcagtt gcagatgttt caccacgcca cgaacgtagt
     1081 tagcccattg atagttttca tgtgcgacaa tgggcgcatc gagggaaaac tcgtcgagct
     1141 gattttcata atcggctgcc atcacgcgaa ctttacggtc atcgcgtggt gcacaactga
     1201 tcacggtttg ataatcaatc gcgcagggca gaacgaaacc gtcgttgtag tcggtgtgtt
     1261 caccaatcaa attcacgcgg ccaggcgcct gaatggtgtg agtggcaggg tagccaaatg
     1321 cgttggcaaa cagagattgt gttttttctt tcagactcat ttcttacact ccggattcgc
     1381 gaaaatggat atcgctgact gcgcgcaaac gctctgctgc ctgttctgcg gtcaggtctc
     1441 gctgggtctc tgccagcatt tcataaccaa ccataaattt acgtacggtg gcggagcgca
     1501 gcagaggcgg ataaaagtgc gcgtgcagcc cttattgccc ctggcgttcc gggttctggt
     1561 caaagctgct gacccggacg tagccgatgc ggtgcccctg cacgatgtct ccttggttga
     1621 aggcggctta agtgcacttt ctgttccgtt gtgcctcaaa gcccatttct gtcaggctga
     1681 aatctataac cttcgcgggc atgtgtcaaa aaatgggaaa gcagactcta ttctgacgaa
     1741 gcggcgcggc cctgcctgac atcaagttag ggtatagcct agattgacat gcgcgatgca
     1801 acccttaact tgcttgcacc tatcgtttcc atgctagccg gctacgtatc gatgaattct
     1861 cgaggcggcc gcgggcccat ggtacccggg gatccggtgc aggccaggct ttttcaggta
     1921 gtcacggccg gtcacgctgc ccactgcaat caccccgccg atatgctcga tcgccggcgc
     1981 cagcgcaacc gggacgatga acagaatggc ctgccagttg aacgcagggg cggtgaaatt
     2041 gggaatttcc agccacggtg cggcagcgat cctggcggtg tcgaccacgc caaaggcgaa
     2101 cgacagggca aagcccacca gcacgccgga gatgatgggt accaggcgga aaatgccctt
     2161 gccgaatacg gcaacgatca aggttgtcag cagcgctggc atcgagatca gcatggccgt
     2221 cttgtacggc atcagtgccg taccgtcccc acctttgccc atcgccatgt tggcggcgat
     2281 cggtgccatg gccaggccga tggagatgat caccgggccg atcaccaccg gcggcagcat
     2341 gcggtcgatg aaaccggtgc ctttgatttt taccatcagc cccatgaagg tgtacacgaa
     2401 gccggcagcc attacgccac ccatggtctc ggcaaggccg aactggcctt tggcgaggat
     2461 gatcggcgtg atgaaggcaa agctcgaggc caggaacacc ggtacctgac gaccggtgac
     2521 cagctggaat agcagcgtgc cgatgccggc ggtgaacagc gcgacgtttg ggtcgaggcc
     2581 ggtgatcagg ggcatcagca ccagcgcgcc gaatgccacg aagagcatct gcgcgcccga
     2641 aacgacctgg cgccagagcg ggtcgttgaa gccgtcctgc atggtcaaag cttcataggg
     2701 gagggctccg aaaggcgggc aaaaaaaccg cgctagatta atctattcag cctgtgctgt
     2761 cgtctggtca ttctggacgt tagtccataa atgcttgatc tgtgacgagc ggatgcgtac
     2821 ctttgcccgc ttttccaaaa tgccccccgg agcgcgccat gtccgccgat ctcgagcata
     2881 tccgtcaagt catgcatgaa gctgactgcc tgtacaccga agccgaagtc gaagcggcca
     2941 tcgccaaagt cggcgagcag atctgcaagg acctgcacga caagaacccg gttgtgttct
     3001 gcgtgatgaa cggtggcctg atcttctccg gcaaactgct gacccacctg cagttcccgc
     3061 tggaagcctc gtacctgcat gctacccgct accgcaacca gaccagcggt ggcgagcttt
     3121 tctggaaggc caagccggaa gtgtcgttca tcgaccgtga cgtgttgatc gttgacgata
     3181 tcctcgacga aggccacacc ctcagcgcca tcatcgagtt ttgcaagcat gccggcgccc
     3241 gctcggtgta caccgctgtg ctgatcgaca aggaccacga ccgcaaggca agccctgacc
     3301 tcaaggccaa ctatgtgggc ctgccttgtg tcgatcgcta tatcttcggt tacggcatgg
     3361 actacaaagg ttactggcgc aacgctaacg gcatcttcgc cgtaaagggg ctgtaaccgt
     3421 gcgccagcct gcgttcatcg accaatcgct gttcgccggg ctggccgaaa aggccgctgc
     3481 caacccgcgc gggcggcagc accataactt tcacgctatg gaagagccct gccaccgcat
     3541 ggccgtcggg ctgcaaccaa gcacctacat tccgccggat ccgggaccaa agcggccatc
     3601 gtgcctcccc actcctgcag ttcgggggca tggatgcgcg gatagccgct gctggtttcc
     3661 tggatgccga cggatttgca ctgccggtag aactccgcga ggtcgtccag cctcaggcag
     3721 cagctgaacc aactcgcgag gggatcgagc ccggggtggg cgaagaactc cagcatgaga
     3781 tccccgcgct ggaggatcat ccagccggcg tcccggaaaa cgattccgaa gcccaacctt
     3841 tcatagaagg cggcggtgga atcgaaatct cgtgatggca ggttgggcgt cgcttggtcg
     3901 gtcatttcga accccagagt cccgctcaga agaactcgtc aagaaggcga tagaaggcga
     3961 tgcgctgcga atcgggagcg gcgataccgt aaagcacgag gaagcggtca gcccattcgc
     4021 cgccaagctc ttcagcaata tcacgggtag ccaacgctat gtcctgatag cggtccgcca
     4081 cacccagccg gccacagtcg atgaatccag aaaagcggcc attttccacc atgatattcg
     4141 gcaagcaggc atcgccatgg gtcacgacga gatcctcgcc gtcgggcatg cgcgccttga
     4201 gcctggcgaa cagttcggct ggcgcgagcc cctgatgctc ttcgtccaga tcatcctgat
     4261 cgacaagacc ggcttccatc cgagtacgtg ctcgctcgat gcgatgtttc gcttggtggt
     4321 cgaatgggca ggtagccgga tcaagcgtat gcagccgccg cattgcatca gccatgatgg
     4381 atactttctc ggcaggagca aggtgagatg acaggagatc ctgccccggc acttcgccca
     4441 atagcagcca gtcccttccc gcttcagtga caacgtcgag cacagctgcg caaggaacgc
     4501 ccgtcgtggc cagccacgat agccgcgctg cctcgtcctg cagttcattc agggcaccgg
     4561 acaggtcggt cttgacaaaa agaaccgggc gcccctgcgc tgacagccgg aacacggcgg
     4621 catcagagca gccgattgtc tgttgtgccc agtcatagcc gaatagcctc tccacccaag
     4681 cggccggaga acctgcgtgc aatccatctt gttcaatcat gcgaaacgat cctcatcctg
     4741 tctcttgatc agatcttgat cccctgcgcc atcagatcct tggcggcaag aaagccatcc
     4801 agtttacttt gcagggcttc ccaaccttac cagagggcgc cccagctggc aattccggtt
     4861 cgcttgctgt ccataaaacc gcccagtcta gctatcgcca tgtaagccca ctgcaagcta
     4921 cctgctttct ctttgcgctt gcgttttccc ttgtccagat agcccagtag ctgacattca
     4981 tccggggtca gcaccgtttc tgcggactgg ctttctacgt gttccgcttc ctttagcagc
     5041 ccttgcgccc tgagtgcttg cggcagcgtg aagcttggcg taatcatggt catagctgtt
     5101 tcctgtgtga aattgttatc cgctcacaat tccacacaac atacgagccg gaagcataaa
     5161 gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca ttaattgcgt tgcgctcact
     5221 gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc
     5281 ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg
     5341 ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc
     5401 cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag
     5461 gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca
     5521 tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca
     5581 ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg
     5641 atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag
     5701 gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt
     5761 tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca
     5821 cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg
     5881 cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa gaacagtatt
     5941 tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc
     6001 cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg
     6061 cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg
     6121 gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta
     6181 gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg
     6241 gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg
     6301 ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc
     6361 atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc cagatttatc
     6421 agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc
     6481 ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag
     6541 tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat
     6601 ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg
     6661 caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt
     6721 gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag
     6781 atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg
     6841 accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt
     6901 aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct
     6961 gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac
     7021 tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat
     7081 aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat
     7141 ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca
     7201 aataggggtt ccgcgcacat ttccccgaaa agtgccacct gacgtcagcc gggcaggata
     7261 ggtgaagtag gcccacccgc gagcgggtgt tccttcttca ctgtcccttc tagaacgtct
     7321 aagaaaccat tattatcatg acattaacct ataaaaatag gcgtatcacg aggccctttc
     7381 gt
//
