LOCUS       sequence-405908-        8811 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Assembly for pyrG deletion in P. digitatum..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8811)
  AUTHORS   Moreno-Gimenez E, Gandia M, Saez Z, Manzanares P, Yenush L, Orzaez 
            D, Marcos JF, Garrigues S
  TITLE     FungalBraid 2.0: expanding the synthetic biology toolbox for the 
            biotechnological exploitation of filamentous fungi.
  JOURNAL   Front Bioeng Biotechnol. 2023 Aug 7;11:1222812. doi: 
            10.3389/fbioe.2023.1222812. eCollection 2023.
  PUBMED    37609115
REFERENCE   2  (bases 1 to 8811)
  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: "Front 
            Bioeng Biotechnol."; date: "2023-08-7"; volume: "11:1222812. doi"; 
            pages: " 10.3389/fbioe.2023.1222812. eCollection 2023"
COMMENT     Sequence Label: FB372
FEATURES             Location/Qualifiers
     source          1..8811
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    2458..2482
                     /label=RB T-DNA repeat
                     /note="right border repeat from nopaline C58 T-DNA"
     CDS             3783..4412
                     /codon_start=1
                     /product="stability protein from the Pseudomonas plasmid 
                     pVS1 (Heeb et al., 2000)"
                     /label=pVS1 StaA
                     /translation="MKVIAVLNQKGGSGKTTIATHLARALQLAGADVLLVDSDPQGSAR
                     DWAAVREDQPLTVVGIDRPTIDRDVKAIGRRDFVVIDGAPQAADLAVSAIKAADFVLIP
                     VQPSPYDIWATADLVELVKQRIEVTDGRLQAAFVVSRAIKGTRIGGEVAEALAGYELPI
                     LESRITQRVSYPGTAAAGTTVLESEPEGDAAREVQALAAEIKSKLI"
     CDS             4841..5914
                     /codon_start=1
                     /product="replication protein from the Pseudomonas plasmid 
                     pVS1 (Heeb et al., 2000)"
                     /label=pVS1 RepA
                     /translation="MSGRKPSGPVQIGAALGDDLVEKLKAAQAAQRQRIEAEARPGESW
                     QAAADRIRKESRQPPAAGAPSIRKPPKGDEQPDFFVPMLYDVGTRDSRSIMDVAVFRLS
                     KRDRRAGEVIRYELPDGHVEVSAGPAGMASVWDYDLVLMAVSHLTESMNRYREGKGDKP
                     GRVFRPHVADVLKFCRRADGGKQKDDLVETCIRLNTTHVAMQRTKKAKNGRLVTVSEGE
                     ALISRYKIVKSETGRPEYIEIELADWMYREITEGKNPDVLTVHPDYFLIDPGIGRFLYR
                     LARRAAGKAEARWLFKTIYERSGSAGEFKKFCFTVRKLIGSNDLPEYDLKEEAGQAGPI
                     LVMRYRNLIEGEASAGS"
     rep_origin      5980..6174
                     /label=pVS1 oriV
                     /note="origin of replication for the Pseudomonas plasmid 
                     pVS1 (Heeb et al., 2000)"
     primer_bind     complement(6410..6432)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    6518..6658
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     6573..6592
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(6673..6690)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6844..7432)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6924..6943)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(7520..8335)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(7520..8335)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     8243..8262
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     misc_feature    8768..8792
                     /label=LB T-DNA repeat
                     /note="left border repeat from nopaline C58 T-DNA"
ORIGIN
        1 aggatgaagg atccagaaaa tccttcaatc tgaagtcatc ctcaatgctc cgaaaggcca
       61 actatatcac aacagtagac aagaaaacct tggaagaccc cggatcggct aaatcgattg
      121 caccgacggg acgtcggaac ctgtgagccc cgaacctcca aatatgtata aaggagccca
      181 ggcatcaggc taaatcacaa cacaaagccg ctctctcact tcatttcccc ggttaatctt
      241 ctcataatgg tcgcattcat tgttcctggc aactacgggt aggttatgcc actcccggct
      301 taaccttcgc gtttcctgtg aaatccgggg ctaacataac ttcccagagc cgtcattgcc
      361 gtggccctcg gtgccatccc tgttttgggc tttattcacg gcaatgtcac tggtagcctg
      421 cgcaaggcgg ccaaggtgcc ctacccccac agctatgcct ccatggagct atgcaaggaa
      481 aacgtgcgta gaaatttttt ttccatctgt cggccttcaa aagcccttca gttcaaggga
      541 aacatgtgta atctctaatg aacaccccat taaacctgct aatctgatta taggcaaaag
      601 ctgagcagtt caactgcgcc cagcgcgccc atgccaactt cctcgagaac gcctcgcaga
      661 caatgctctt taccttggtc gccggtctaa agtatccgga gtgggccgct ggtctgggcg
      721 ctctctgggt tttcttccgc gttctcttcc tctacggcta cgtttactcg ggcaagccgc
      781 agggtaaggg ccgcttcatg ggcgggttct tctggcttgt gcagggtgct ctttggggat
      841 tgagcatcta cggtgttgga aggcctctgc tttctttcta agagcttgaa gcctggggat
      901 cctctaaaat gtcttgaaat tgaagatgta ctctgtattt ataagtccaa tgcaagcaac
      961 gtattagtta tgcgccacag ttccagcctc ggaaggttat gttgtaggcc tgcggggccc
     1021 accgtggcag aaaaagttcc ccgatggtct tcaaatcgaa cttgctcctc atcttctgct
     1081 ttttggaccc caaaaagcct atagttctct gaaggcattt gtcgtttgat tcatttccat
     1141 cctttcgacc cctcaaaaca caagctaaat acacaacaat cgcctcttca ccccgccagc
     1201 cccgccaatg ccagcccggt actttcttgc ctcagattat tagtgagtct ttccagggct
     1261 ctaaaagggc ttccctgcat gtgtaaacga tgcaagggca atttcaagtc ttacgtgtgc
     1321 tcgacgatat ggttcactcc cccttcattg aatcatcaaa tcaaacgcga atctctgaat
     1381 attcgaatgt tgggagtagc ttgataaggc ggctccaaac ttcagatgtc gagataccgg
     1441 acaacagatc agagtaatac tgtaattatg tgtcaacctc atcagaacca gcagaactag
     1501 aagcgtaagc atgtcgacat atagaccgga taatgcctgg cactcgagat ctttcccttt
     1561 ccgttttagt gctcacccct tgccgctttt tgatcactga gtaaaaccga gcacgtccct
     1621 gagtatcctg gcttgatcac aggtcattac ataatcaact gatctgctca ctgagcatca
     1681 tctgagcact ctgatttgat cactgagtgc tcactgagtg tagggatgag cgggctgagt
     1741 aagctacatc aataaaatgc gacacgctaa tcccatcacg tgcaaccatt cccattcaga
     1801 tagcccatca aaaacggact ggcacaatca atgtgcccgt tcgttctagc ggcctcgatt
     1861 ctatccaagt aaaaaagccc cacagtgggg ctattaattg acctaccatc ttctcatttc
     1921 agcctatgct cctgggaata cctcacggaa taccctaatc gggctttggg ctactaccag
     1981 ggcgagggcg attgaatgag tttccttgaa gatctagaat cattccctgt gtcggcaaca
     2041 ggagaaatta gtgttcagta agtgacgatg atgagccact tctgtcgcca atgagcgcac
     2101 gacccaaagg atctcctggg cagataacgt catgtctttc acgccattgc taagtaatct
     2161 gtaagccgaa gacaaaccat caataataat acccgatgag ctcctgggag cctgccatgc
     2221 attcccatga gcccactaag ctgtattacc agtgtcatag gggcccatag ctatttcgtt
     2281 tgaatcctag gaactctaaa cattgccccc cacccctagc ctgtctgatt tctgcctaaa
     2341 gcgatgccat tccaaaaggg atatgtatct agccacttgc tcggaaacaa tggtcttcat
     2401 cattcctctg cctgcttttg ctaaccattt gggacaccgc tgtcatgaga cgaattctga
     2461 caggatatat tggcgggtaa acctaagaga aaagagcgtt tattagaata atcggatatt
     2521 taaaagggcg tgaaaaggtt tatccgttcg tccatttgta tgtgcatgcc aaccacaggg
     2581 ttcccctcgg gatcaaagta ctttgatcca acccctccgc tgctatagtg cagtcggctt
     2641 ctgacgttca gtgcagccgt catctgaaaa cgacatgtcg cacaagtcct aagttacgcg
     2701 acaggctgcc gccctgccct tttcctggcg ttttcttgtc gcgtgtttta gtcgcataaa
     2761 gtagaatact tgcgactaga accggagaca ttacgccatg aacaagagcg ccgccgctgg
     2821 cctgctgggc tatgcccgcg tcagcaccga cgaccaggac ttgaccaacc aacgggccga
     2881 actgcacgcg gccggctgca ccaagctgtt ttccgagaag atcaccggca ccaggcgcga
     2941 ccgcccggag ctggccagga tgcttgacca cctacgccct ggcgacgttg tgacagtgac
     3001 caggctagac cgcctggccc gcagcacccg cgacctactg gacattgccg agcgcatcca
     3061 ggaggccggc gcgggcctgc gtagcctggc agagccgtgg gccgacacca ccacgccggc
     3121 cggccgcatg gtgttgaccg tgttcgccgg cattgccgag ttcgagcgtt ccctaatcat
     3181 cgaccgcacc cggagcgggc gcgaggccgc caaggcccga ggcgtgaagt ttggcccccg
     3241 ccctaccctc accccggcac agatcgcgca cgcccgcgag ctgatcgacc aggaaggccg
     3301 caccgtgaaa gaggcggctg cactgcttgg cgtgcatcgc tcgaccctgt accgcgcact
     3361 tgagcgcagc gaggaagtga cgcccaccga ggccaggcgg cgcggtgcct tccgtgagga
     3421 cgcattgacc gaggccgacg ccctggcggc cgccgagaat gaacgccaag aggaacaagc
     3481 atgaaaccgc accaggacgg ccaggacgaa ccgtttttca ttaccgaaga gatcgaggcg
     3541 gagatgatcg cggccgggta cgtgttcgag ccgcccgcgc acctctcaac cgtgcggctg
     3601 catgaaatcc tggccggttt gtctgatgcc aagctggcgg cctggccggc cagcttggcc
     3661 gctgaagaaa ccgagcgccg ccgtctaaaa aggtgatgtg tatttgagta aaacagcttg
     3721 cgtcatgcgg tcgctgcgta tatgatgcga tgagtaaata aacaaatacg caaggggaac
     3781 gcatgaaggt tatcgctgta cttaaccaga aaggcgggtc aggcaagacg accatcgcaa
     3841 cccatctagc ccgcgccctg caactcgccg gggccgatgt tctgttagtc gattccgatc
     3901 cccagggcag tgcccgcgat tgggcggccg tgcgggaaga tcaaccgcta accgttgtcg
     3961 gcatcgaccg cccgacgatt gaccgcgacg tgaaggccat cggccggcgc gacttcgtag
     4021 tgatcgacgg agcgccccag gcggcggact tggctgtgtc cgcgatcaag gcagccgact
     4081 tcgtgctgat tccggtgcag ccaagccctt acgacatatg ggccaccgcc gacctggtgg
     4141 agctggttaa gcagcgcatt gaggtcacgg atggaaggct acaagcggcc tttgtcgtgt
     4201 cgcgggcgat caaaggcacg cgcatcggcg gtgaggttgc cgaggcgctg gccgggtacg
     4261 agctgcccat tcttgagtcc cgtatcacgc agcgcgtgag ctacccaggc actgccgccg
     4321 ccggcacaac cgttcttgaa tcagaacccg agggcgacgc tgcccgcgag gtccaggcgc
     4381 tggccgctga aattaaatca aaactcattt gagttaatga ggtaaagaga aaatgagcaa
     4441 aagcacaaac acgctaagtg ccggccgtcc gagcgcacgc agcagcaagg ctgcaacgtt
     4501 ggccagcctg gcagacacgc cagccatgaa gcgggtcaac tttcagttgc cggcggagga
     4561 tcacaccaag ctgaagatgt acgcggtacg ccaaggcaag accattaccg agctgctatc
     4621 tgaatacatc gcgcagctac cagagtaaat gagcaaatga ataaatgagt agatgaattt
     4681 tagcggctaa aggaggcggc atggaaaatc aagaacaacc aggcaccgac gccgtggaat
     4741 gccccatgtg tggaggaacg ggcggttggc caggcgtaag cggctgggtt gcctgccggc
     4801 cctgcaatgg cactggaacc cccaagcccg aggaatcggc gtgagcggtc gcaaaccatc
     4861 cggcccggta caaatcggcg cggcgctggg tgatgacctg gtggagaagt tgaaggccgc
     4921 gcaggccgcc cagcggcaac gcatcgaggc agaagcacgc cccggtgaat cgtggcaagc
     4981 ggccgctgat cgaatccgca aagaatcccg gcaaccgccg gcagccggtg cgccgtcgat
     5041 taggaagccg cccaagggcg acgagcaacc agattttttc gttccgatgc tctatgacgt
     5101 gggcacccgc gatagtcgca gcatcatgga cgtggccgtt ttccgtctgt cgaagcgtga
     5161 ccgacgagct ggcgaggtga tccgctacga gcttccagac gggcacgtag aggtttccgc
     5221 agggccggcc ggcatggcca gtgtgtggga ttacgacctg gtactgatgg cggtttccca
     5281 tctaaccgaa tccatgaacc gataccggga agggaaggga gacaagcccg gccgcgtgtt
     5341 ccgtccacac gttgcggacg tactcaagtt ctgccggcga gccgatggcg gaaagcagaa
     5401 agacgacctg gtagaaacct gcattcggtt aaacaccacg cacgttgcca tgcagcgtac
     5461 gaagaaggcc aagaacggcc gcctggtgac ggtatccgag ggtgaagcct tgattagccg
     5521 ctacaagatc gtaaagagcg aaaccgggcg gccggagtac atcgagatcg agctagctga
     5581 ttggatgtac cgcgagatca cagaaggcaa gaacccggac gtgctgacgg ttcaccccga
     5641 ttactttttg atcgatcccg gcatcggccg ttttctctac cgcctggcac gccgcgccgc
     5701 aggcaaggca gaagccagat ggttgttcaa gacgatctac gaacgcagtg gcagcgccgg
     5761 agagttcaag aagttctgtt tcaccgtgcg caagctgatc gggtcaaatg acctgccgga
     5821 gtacgatttg aaggaggagg cggggcaggc tggcccgatc ctagtcatgc gctaccgcaa
     5881 cctgatcgag ggcgaagcat ccgccggttc ctaatgtacg gagcagatgc tagggcaaat
     5941 tgccctagca ggggaaaaag gtcgaaaagg actctttcct gtggatagca cgtacattgg
     6001 gaacccaaag ccgtacattg ggaaccggaa cccgtacatt gggaacccaa agccgtacat
     6061 tgggaaccgg tcacacatgt aagtgactga tataaaagag aaaaaaggcg atttttccgc
     6121 ctaaaactct ttaaaactta ttaaaactct taaaacccgc ctggcctgtg cataactgtc
     6181 tggccagcgc acagccgaag agctgcaaaa agcgcctacc cttcggtcgc tgcgctccct
     6241 acgccccgcc gcttcgcgtc ggcctatcgc ggccgctggc cgctcaaaaa tggctggcct
     6301 acggccaggc aatctaccag ggcgcggaca agccgcgccg tcgccactcg accgccggcg
     6361 cccacatcaa ggcaccctgc ctcgcgcgtt tcggtgatga cggtgaaaac ctctgacaca
     6421 tgcagctccc ggtgacggtc acagcttgtc tgtaagcgga tgccgggagc agacaagccc
     6481 gtcagggcgc gtcagcgggt gttggcgggt gtcggggcgc agccatgacc cagtcacgta
     6541 gcgatagcgg agtgtatact ggcttaacta tgcggcatca gagcagattg tactgagagt
     6601 gcaccatatg cggtgtgaaa taccgcacag atgcgtaagg agaaaatacc gcatcaggcg
     6661 ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt
     6721 atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa
     6781 gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc
     6841 gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag
     6901 gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt
     6961 gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg
     7021 aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg
     7081 ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg
     7141 taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac
     7201 tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg
     7261 gcctaactac ggctacacta gaaggacagt atttggtatc tgcgctctgc tgaagccagt
     7321 taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg
     7381 tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc
     7441 tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt
     7501 ggtcatgcat tctaggtgat tagaaaaact catcgagcat caaatgaaac tgcaatttat
     7561 tcatatcagg attatcaata ccatattttt gaaaaagccg tttctgtaat gaaggagaaa
     7621 actcaccgag gcagttccat aggatggcaa gatcctggta tcggtctgcg attccgactc
     7681 gtccaacatc aatacaacct attaatttcc cctcgtcaaa aataaggtta tcaagtgaga
     7741 aatcaccatg agtgacgact gaatccggtg agaatggcaa aagtttatgc atttctttcc
     7801 agacttgttc aacaggccag ccattacgct cgtcatcaaa atcactcgca tcaaccaaac
     7861 cgttattcat tcgtgattgc gcctgagcga gtcgaaatac gcgatcgctg ttaaaaggac
     7921 aattacaaac aggaatcgaa tgcaaccggc gcaggaacac tgccagcgca tcaacaatat
     7981 tttcacctga atcaggatat tcttctaata cctggaatgc tgttttccct gggatcgcag
     8041 tggtgagtaa ccatgcatca tcaggagtac ggataaaatg cttgatggtc ggaagaggca
     8101 taaattccgt cagccagttt agtctgacca tctcatctgt aacatcattg gcaacgctac
     8161 ctttgccatg tttcagaaac aactctggcg catcgggctt cccatacaat cggtagattg
     8221 tcgcacctga ttgcccgaca ttatcgcgag cccatttata cccatataaa tcagcatcca
     8281 tgttggaatt taatcgcggc cttgagcaag acgtttcccg ttgaatatgg ctcataacag
     8341 aacttattat ttccttcctc ttttctacag tatttaaaga taccccaaga agctaattat
     8401 aacaagacga actccaattc actgttcctt gcattctaaa accttaaata ccagaaaaca
     8461 gctttttcaa agttgttttc aaagttggcg tataacatag tatcgacgga gccgattttg
     8521 aaaccgcggt gatcacaggc agcaacgctc tgtcatcgtt acaatcaaca tgctaccctc
     8581 cgcgagatca tccgtgtttc aaacccggca gcttagttgc cgttcttccg aatagcatcg
     8641 gtaacatgag caaagtctgc cgccttacaa cggctctccc gctgacgccg tcccggactg
     8701 atgggctgcc tgtatcgagt ggtgattttg tgccgagctg ccggtcgggg agctgttggc
     8761 tggctggtgg caggatatat tgtggtgtaa acataacgaa ttcgtctcag g
//
