LOCUS       sequence-431357-        4506 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  deletion of amaB.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4506)
  AUTHORS   Ruhl IA, Woodworth SP, Haugen SJ, Alt HM, Beckham GT, Johnson CW
  TITLE     Production of Vanillin From Ferulic Acid by Pseudomonas putida 
            KT2440 Using Metabolic Engineering and In Situ Product Recovery.
  JOURNAL   Microb Biotechnol. 2025 May;18(5):e70152. doi: 
            10.1111/1751-7915.70152.
  PUBMED    40410945
REFERENCE   2  (bases 1 to 4506)
  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: "Microb 
            Biotechnol."; date: "2025-05"; volume: "18(5):e70152. doi"; pages: "
            10.1111/1751-7915.70152"
COMMENT     Sequence Label: pIAR008
FEATURES             Location/Qualifiers
     source          1..4506
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     68..87
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     CDS             join(205..291,292..1626)
                     /codon_start=1
                     /gene="Bacillus subtilis sacB"
                     /product="secreted levansucrase that renders bacterial 
                     growth sensitive to sucrose"
                     /label=SacB
                     /note="negative selection marker"
                     /translation="MNIKKFAKQATVLTFTTALLAGGATQAFAKETNQKPYKETYGISH
                     ITRHDMLQIPEQQKNEKYQVSEFDSSTIKNISSAKGLDVWDSWPLQNADGTVANYHGYH
                     IVFALAGDPKNADDTSIYMFYQKVGETSIDSWKNAGRVFKDSDKFDANDSILKDQTQEW
                     SGSATFTSDGKIRLFYTDFSGKHYGKQTLTTAQVNVSASDSSLNINGVEDYKSIFDGDG
                     KTYQNVQQFIDEGNYSSGDNHTLRDPHYVEDKGHKYLVFEANTGTEDGYQGEESLFNKA
                     YYGKSTSFFRQESQKLLQSDKKRTAELANGALGMIELNDDYTLKKVMKPLIASNTVTDE
                     IERANVFKMNGKWYLFTDSRGSKMTIDGITSNDIYMLGYVSNSLTGPYKPLNKTGLVLK
                     MDLDPNDVTFTYSHFAVPQAKGNNVVITSYMTNRGFYADKQSTFAPSFLLNIKGKKTSV
                     VKDSILEQGQLTVNK"
     rep_origin      1659..2247
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2148..2167
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2259..2275
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2259..2275
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3812..3829)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3812..3828)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             join(3910..4506,1..198)
                     /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(3964..3983)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
ORIGIN
        1 aatggccgct tttctggatt catcgactgt ggccggctgg gtgtggcgga ccgctatcag
       61 gacatagcgt tggctacccg tgatattgct gaagagcttg gcggcgaatg ggctgaccgc
      121 ttcctcgtgc tttacggtat cgccgctccc gattcgcagc gcatcgcctt ctatcgcctt
      181 cttgacgagt tcttctgagc gacgatgaac atcaaaaagt ttgcaaaaca agcaacagta
      241 ttaaccttta ctaccgcact gctggcagga ggcgcaactc aagcgtttgc gaaagaaacg
      301 aaccaaaagc catataagga aacatacggc atttcccata ttacacgcca tgatatgctg
      361 caaatccctg aacagcaaaa aaatgaaaaa tatcaagttt ctgaatttga ttcgtccaca
      421 attaaaaata tctcttctgc aaaaggcctg gacgtttggg acagctggcc attacaaaac
      481 gctgacggca ctgtcgcaaa ctatcacggc taccacatcg tctttgcatt agccggagat
      541 cctaaaaatg cggatgacac atcgatttac atgttctatc aaaaagtcgg cgaaacttct
      601 attgacagct ggaaaaacgc tggccgcgtc tttaaagaca gcgacaaatt cgatgcaaat
      661 gattctatcc taaaagacca aacacaagaa tggtcaggtt cagccacatt tacatctgac
      721 ggaaaaatcc gtttattcta cactgatttc tccggtaaac attacggcaa acaaacactg
      781 acaactgcac aagttaacgt atcagcatca gacagctctt tgaacatcaa cggtgtagag
      841 gattataaat caatctttga cggtgacgga aaaacgtatc aaaatgtaca gcagttcatc
      901 gatgaaggca actacagctc aggcgacaac catacgctga gagatcctca ctacgtagaa
      961 gataaaggcc acaaatactt agtatttgaa gcaaacactg gaactgaaga tggctaccaa
     1021 ggcgaagaat ctttatttaa caaagcatac tatggcaaaa gcacatcatt cttccgtcaa
     1081 gaaagtcaaa aacttctgca aagcgataaa aaacgcacgg ctgagttagc aaacggcgct
     1141 ctcggtatga ttgagctaaa cgatgattac acactgaaaa aagtgatgaa accgctgatt
     1201 gcatctaaca cagtaacaga tgaaattgaa cgcgcgaacg tctttaaaat gaacggcaaa
     1261 tggtacctgt tcactgactc ccgcggatca aaaatgacga ttgacggcat tacgtctaac
     1321 gatatttaca tgcttggtta tgtttctaat tctttaactg gcccatacaa gccgctgaac
     1381 aaaactggcc ttgtgttaaa aatggatctt gatcctaacg atgtaacctt tacttactca
     1441 cacttcgctg tacctcaagc gaaaggaaac aatgtcgtga ttacaagcta tatgacaaac
     1501 agaggattct acgcagacaa acaatcaacg tttgcgccga gcttcctgct gaacatcaaa
     1561 ggcaagaaaa catctgttgt caaagacagc atccttgaac aaggacaatt aacagttaac
     1621 aaataatcag accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc
     1681 gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat
     1741 caagagctac caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat
     1801 actgttcttc tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct
     1861 acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt
     1921 cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg
     1981 gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta
     2041 cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg
     2101 gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg
     2161 tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc
     2221 tcgtcagggg ggcggagcct atggaaaaac gcctcacaca ggaaacagct atgacatgat
     2281 tacgaattca ggcgcgttac agcagccgcc aggccctggg tgtagcccat ggggttgatg
     2341 gtgccagcac ggcgatcaag cagcgcagcg gaaatcttgt cggtaccgca gtattcctgg
     2401 cactgggcac cggtcagcag ctcgacgtcg gcacctcggc ggcgccattg ttcgtgacgc
     2461 gcttcgaggt cggcgatacc ggtggcgttg tgcgccatgt gcagcgtgcc tttgtgctgg
     2521 gcctggcagt cgatgccgag gcgctcgatc atggcgaaca cttcagccgg ggcttcgcca
     2581 agcaccttgt tcaggcggct accctgcttc tggccaaggg tggcctcgac gtcgtcgggg
     2641 cggatccacg tgccggcgtt gaccaggccg acgttgcgcc cggagccacc gtggccgatc
     2701 ttccaggcct ccagcacgat caccgacttg ccctgttcga gcaggtggat ggccgccgac
     2761 aggccggtaa taccgccgcc gatcacacag acatcggcct tgtgctcacc ggccagggcc
     2821 tgggcggcga cggtcggttt gctgacgtgt tcccacaagc attgttgacg cagttcggac
     2881 atggccggct ccagcaattt gttcttgttg gggccgcagg gcggcccttt cgcgggtaaa
     2941 cccgctccta caggggtcgg tgtcgcttgg acgaaccctg taggagcggg tctacccgcg
     3001 aaccgggggc caggcccccg gccatgcagc atcaatacag atatcttgtg catctcctac
     3061 atcggtgatg gggtggaaaa atgatgtgga tcagtattaa tccgatcaca taggtacaac
     3121 aaacgacctt tattccgcat atcattccgt caggtaatga tttgagccgc agagaccgct
     3181 atgtcaaaac gcctggtacc ctccatgacc gccctgcagt gctttgaagc tgcagcccgc
     3241 catctgagtt ttacccgtgc cgccgaagaa ctgcacctga cccagagcgc ggtcagcaag
     3301 caggtggcgc agctggaaga catgctgcgc caccacctgt tcctgcgcat ccgccggcgc
     3361 ttgcagctga ccccggccgg cagcctgtac ctggccgagg tcaacaaaat cctgacccag
     3421 gtggatatgt ccagccgcta tgtgctcacc tacggcgagc agaccgagat attgaaggta
     3481 gccactcaac cgagttttgg cgtgcgctgg ttgatcccgc acctcaaggg gtttggcaaa
     3541 cggcacggca atatccacct ggacattcgc aacgagatgg agccgttcgc cctgctgcaa
     3601 ggctcggctg acgtggtgtt cttctatggc cagggcacct ggccgggggc cacttgcgtg
     3661 gagttgtttc gcgaggaggt ggtgccggtg tgtgcgccag agctgctggc cgggcgcaag
     3721 ctggccggtg ccgaggagct ggccgagctg gtgctgctgc aaagcacctc gcgcccggag
     3781 gcctggcatg agtggttcct ggaagcttgg cactggccgt cgttttacaa cgtcgtgacc
     3841 ggaattgcca gctggggcgc cctctggtaa ggttgggaag ccctgcaaac aggatgagga
     3901 tcgtttcgca tgattgaaca agatggattg cacgcaggtt ctccggccgc ttgggtggag
     3961 aggctattcg gctatgactg ggcacaacag acaatcggct gctctgatgc cgccgtgttc
     4021 cggctgtcag cgcaggggcg cccggttctt tttgtcaaga ccgacctgtc cggtgccctg
     4081 aatgaactcc aagacgaggc agcgcggcta tcgtggctgg ccacgacggg cgttccttgc
     4141 gcagctgtgc tcgacgttgt cactgaagcg ggaagggact ggctgctatt gggcgaagtg
     4201 ccggggcagg atctcctgtc atctcacctt gctcctgccg agaaagtatc catcatggct
     4261 gatgcaatgc ggcggctgca tacgcttgat ccggctacct gcccattcga ccaccaagcg
     4321 aaacatcgca tcgagcgagc acgtactcgg atggaagccg gtcttgtcga tcaggatgat
     4381 ctggacgaag agcatcaggg gctcgcgcca gccgaactgt tcgccaggct caaggcgcgg
     4441 atgcccgacg gcgaggatct cgtcgtgacc catggcgatg cctgcttgcc gaatatcatg
     4501 gtggaa
//
