LOCUS       sequence-424009-        4494 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  deletion of areA(calA).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4494)
  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 4494)
  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: pIAR004
FEATURES             Location/Qualifiers
     source          1..4494
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      1..589
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     490..509
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     601..617
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     601..617
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2142..2159)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2142..2158)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             2240..3034
                     /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(2294..2313)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2904..2923
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     CDS             join(3041..3127,3128..4462)
                     /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"
ORIGIN
        1 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
       61 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
      121 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
      181 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
      241 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
      301 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
      361 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
      421 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
      481 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
      541 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgcctcaca
      601 caggaaacag ctatgacatg attacgaatt cccctgttgc tccaccagtg cccgcaacac
      661 accggcctgc gggccgcgca gcgcggtata cagcacctcg cgtacgcgcg ccgggcccag
      721 taccttggcg tcaagcgggt cctgcaggca gtgcagcagc cgttccacac tttcgcgcat
      781 gggtgcatcc agtgccgccg aggtcatcga ttgcggggtt tgcgcctgca cttcagcgtc
      841 cggtgccaag cccatgctct gcaccaactc gcccagcacg ccacggtcga tgcccaccgc
      901 cacgcccaac agcggcgcct cgcgggtggc aaaggtttca cacatgaacg gcaccgacaa
      961 cgcctgcacc aggtaatgcc ctgcaccgta ctccagggta cgcggcccca ggcgtgccag
     1021 cttgctgccc tgggccacga tcatgaggct gggttcgtaa atctgtgggc tgctggccac
     1081 atagtggtcc cagctcaaca cctgcacttg gggcaggagg gtgggcacga agcccgggcg
     1141 cgtggtgagg ctgcggatca gtgagcagag cggcgcattg gcatcgacgt gacgggtcag
     1201 ttgcatggtg gaccaagggc agagaatcag acagcttgat catcgcagat tgagggcatg
     1261 gcgccagcca tggcttggca tcttcgtaga atcaggcatg catgccggag aatcagtcgt
     1321 gggcgaggcg gcgaacgacg cgcaaaatgt gtcgcctgaa tcgtttcact tcactgcgag
     1381 ggatatcttg atcggttgcc tgtacgggcc tcttcgcggg caagcccgct cctacacagg
     1441 ggtgatgtag ggcttgcgcg cggtggggcc gtacaggcag caacgatgca gccgttttac
     1501 caggccattg gcttgtccca gcccttccag aacaaccatc gccgcacctc gccgtgcgcc
     1561 ccggtaccga tctgccactc gggccgcccg ctatccagcg caatcaccga aataaccccc
     1621 gcatggctcg ccgccaccag cgtttgcccg tcggcgctga tatccatcgc actgatggtc
     1681 gagcccaggt aatgctgcca atgctccgta ccctcttcgc caaacgcccg caggtagcca
     1741 taggcatcac ccacaatgta ctcgccatcg cgcgctaccc ccgcatacac ccttgcgcca
     1801 tcctgcacca gcggggtacg cgggtcctgg ctgtagtaat cggtatccag cccgtgcagg
     1861 tctgccaccc tcacggccag cgtggcaccg ctgtagaaat ggcaggcatt gacgatcaac
     1921 tggtcaccgg cgcggttgaa caaggcaaag tgtgggtatt cactgcccgg gccgatgtgc
     1981 gccagcggct tgagtcgttc gtccagaaca agatgtcggc tgccttgctc acccacgacg
     2041 atcagccggc catccgccga caccgcacca tgctccatgc tcagccccag gctgatgtcg
     2101 tccgggtcgg tgccctcggc cagctcttcg agaagcttgg cactggccgt cgttttacaa
     2161 cgtcgtgacc ggaattgcca gctggggcgc cctctggtaa ggttgggaag ccctgcaaac
     2221 aggatgagga tcgtttcgca tgattgaaca agatggattg cacgcaggtt ctccggccgc
     2281 ttgggtggag aggctattcg gctatgactg ggcacaacag acaatcggct gctctgatgc
     2341 cgccgtgttc cggctgtcag cgcaggggcg cccggttctt tttgtcaaga ccgacctgtc
     2401 cggtgccctg aatgaactcc aagacgaggc agcgcggcta tcgtggctgg ccacgacggg
     2461 cgttccttgc gcagctgtgc tcgacgttgt cactgaagcg ggaagggact ggctgctatt
     2521 gggcgaagtg ccggggcagg atctcctgtc atctcacctt gctcctgccg agaaagtatc
     2581 catcatggct gatgcaatgc ggcggctgca tacgcttgat ccggctacct gcccattcga
     2641 ccaccaagcg aaacatcgca tcgagcgagc acgtactcgg atggaagccg gtcttgtcga
     2701 tcaggatgat ctggacgaag agcatcaggg gctcgcgcca gccgaactgt tcgccaggct
     2761 caaggcgcgg atgcccgacg gcgaggatct cgtcgtgacc catggcgatg cctgcttgcc
     2821 gaatatcatg gtggaaaatg gccgcttttc tggattcatc gactgtggcc ggctgggtgt
     2881 ggcggaccgc tatcaggaca tagcgttggc tacccgtgat attgctgaag agcttggcgg
     2941 cgaatgggct gaccgcttcc tcgtgcttta cggtatcgcc gctcccgatt cgcagcgcat
     3001 cgccttctat cgccttcttg acgagttctt ctgagcgacg atgaacatca aaaagtttgc
     3061 aaaacaagca acagtattaa cctttactac cgcactgctg gcaggaggcg caactcaagc
     3121 gtttgcgaaa gaaacgaacc aaaagccata taaggaaaca tacggcattt cccatattac
     3181 acgccatgat atgctgcaaa tccctgaaca gcaaaaaaat gaaaaatatc aagtttctga
     3241 atttgattcg tccacaatta aaaatatctc ttctgcaaaa ggcctggacg tttgggacag
     3301 ctggccatta caaaacgctg acggcactgt cgcaaactat cacggctacc acatcgtctt
     3361 tgcattagcc ggagatccta aaaatgcgga tgacacatcg atttacatgt tctatcaaaa
     3421 agtcggcgaa acttctattg acagctggaa aaacgctggc cgcgtcttta aagacagcga
     3481 caaattcgat gcaaatgatt ctatcctaaa agaccaaaca caagaatggt caggttcagc
     3541 cacatttaca tctgacggaa aaatccgttt attctacact gatttctccg gtaaacatta
     3601 cggcaaacaa acactgacaa ctgcacaagt taacgtatca gcatcagaca gctctttgaa
     3661 catcaacggt gtagaggatt ataaatcaat ctttgacggt gacggaaaaa cgtatcaaaa
     3721 tgtacagcag ttcatcgatg aaggcaacta cagctcaggc gacaaccata cgctgagaga
     3781 tcctcactac gtagaagata aaggccacaa atacttagta tttgaagcaa acactggaac
     3841 tgaagatggc taccaaggcg aagaatcttt atttaacaaa gcatactatg gcaaaagcac
     3901 atcattcttc cgtcaagaaa gtcaaaaact tctgcaaagc gataaaaaac gcacggctga
     3961 gttagcaaac ggcgctctcg gtatgattga gctaaacgat gattacacac tgaaaaaagt
     4021 gatgaaaccg ctgattgcat ctaacacagt aacagatgaa attgaacgcg cgaacgtctt
     4081 taaaatgaac ggcaaatggt acctgttcac tgactcccgc ggatcaaaaa tgacgattga
     4141 cggcattacg tctaacgata tttacatgct tggttatgtt tctaattctt taactggccc
     4201 atacaagccg ctgaacaaaa ctggccttgt gttaaaaatg gatcttgatc ctaacgatgt
     4261 aacctttact tactcacact tcgctgtacc tcaagcgaaa ggaaacaatg tcgtgattac
     4321 aagctatatg acaaacagag gattctacgc agacaaacaa tcaacgtttg cgccgagctt
     4381 cctgctgaac atcaaaggca agaaaacatc tgttgtcaaa gacagcatcc ttgaacaagg
     4441 acaattaaca gttaacaaat aatcagaccc cgtagaaaag atcaaaggat cttc
//
