LOCUS       sequence-193900-        5350 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  BB3 recipient for homologous integration in pyrG locus with linker 
            AF.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5350)
  AUTHORS   Sarkari P, Marx H, Blumhoff ML, Mattanovich D, Sauer M, Steiger MG
  TITLE     An efficient tool for metabolic pathway construction and gene 
            integration for Aspergillus niger.
  JOURNAL   Bioresour Technol. 2017 May 4. pii: S0960-8524(17)30643-0. doi: 
            10.1016/j.biortech.2017.05.004.
  PUBMED    28533066
REFERENCE   2  (bases 1 to 5350)
  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: "Bioresour 
            Technol. 2017 May 4. pii: S0960-8524(17)30643-0. doi: 
            10.1016/j.biortech.2017.05.004."
COMMENT     Sequence Label: pMST638_BB3_L_AF_hyg_KanOri_pyrG5
FEATURES             Location/Qualifiers
     source          1..5350
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     264..281
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     265..281
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     regulatory      1630..1639
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2222..3247
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     CDS             2222..3247
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     primer_bind     complement(3910..3926)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3910..3926)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     terminator      3974..4005
                     /label=tonB terminator
                     /note="bidirectional E. coli tonB-P14 transcription 
                     terminator"
     promoter        4006..4108
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     CDS             4109..4924
                     /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="MSHIQRETSCLKPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNGDRVFRLAQAQSRMNNGLVGA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(4182..4201)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     terminator      4948..4975
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     rep_origin      complement(join(4987..5350,1..224))
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5067..5086)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac
       61 tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc agttacctcg
      121 gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt
      181 ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgattt
      241 tctaccgaag aaaggcccac ccgtgtaaaa cgacggccag tttatctagt cagcttgatt
      301 ctagctgatc gtggaccgga aggtgagcca gtgagttgat tgcagtccag ttacgctgga
      361 gtctgaggct cgtcctgaat gatatgcggc ccgctgtagg tcaattgcga cttggaggaa
      421 gggatccgcc aaggatgctc tggatccgca tccggccgct cttgcgcccc atcaatcgcc
      481 cgactataaa tcgaactact ttcggcatct tctagacttc ctaataccgc ctagtcatag
      541 cagattcaag ctgagaacac cacaagtaaa tatcacccat catgcttacc ctgaccgtcc
      601 ctgaaaacta cgggtatgtg ccaattctac aattccttgc agacaatgcc attctcccca
      661 tgaagtctga tgctaactat cctgcagctc tgtcattgcc gtcgctctgg gtgccatccc
      721 cgtcctgagc ttcgtccatg gcgccgtcgt gtctcgtctc cgcaaggaag ctgattgccc
      781 ctaccctcac tgctatgcga ccgtagagca gtgcaagacc aacgtaagcc aacctcacac
      841 aaacaggatt cctcgagcta acatacattc cgaaccgtgc agcccaaggc cgagcagttc
      901 aactgcgctc agcgcgctca tgccaacttc cttgagaact ccagccaaac tatgctcttc
      961 ctcctggtag ctggactgaa gtacccccag ttggcgactg gcctcggaag catctgggtc
     1021 ctcggtcgct cactgttcct ttacggatat gtgtactccg gcaagccgcg gggtcgcggt
     1081 cgtttgtacg gcagcttcta cttgcttgca cagggagctc tctggggctt gacgtctttt
     1141 ggagttgcga gggagttgat ttcctacttc taagtttgga ctgaatccgt ggtgtgattg
     1201 aggtgattgg cgatgtttgg ctataccagc tatatgtaat aatctctact gtatactact
     1261 attcaacgca ttttactatg cgtgctgcta gggtcggcaa tgacaatggc aatctgactg
     1321 acgtggtcta tttctccatg tgcagcaggg aatacgagct ccaatggacc tcgggagtgg
     1381 cacagtcaat ggcaaggaaa ctggagcgca aaaaaccccg cccctgacag ggcggggttt
     1441 tttcgcgatc ggagaccgga tccctcgagg gtctccgctt catgaataac ggtgagacta
     1501 gcggccggtc cccttatccc agctgttcca cgttggcctg cccctcagtt agcgctcaac
     1561 tcaatgcccc tcactggcga ggcgagggca aggatggagg ggcagcatcg cctgagttgg
     1621 agcaaagcgg ccgccatggg agcagcgaac caacggaggg atgccgtgct ttgtcgtggc
     1681 tgctgtggcc aatccgggcc cttggttggc tcacagagcg ttgctgtgag tccatgagct
     1741 attattgcta ggtacagtat agagagagga gagagagaga gagagagaga ggggaaaaaa
     1801 ggtgaggttg aagtgagaaa aaaaaaaaaa aaatccaacc actgacggct gccggctctg
     1861 ccacccccct ccctccaccc cagaccacct gcacactcag cgcgcagcat cacctaatct
     1921 tggctcgcct tcccgcagct caggttgttt tttttttctc tctccctcgt cgaagccgcc
     1981 cttgttccct tatttatttc cctctccatc cttgtctgcc tttggtccat ctgccccttt
     2041 gtctgcatct cttttgcacg catcgcctta tcgtcgtctc ttttttcact cacgggagct
     2101 tgacgatgac ctgactcgtg agcctcacct gctgatttct ctccccccct cccgaccggc
     2161 ttgacttttg tttctcctcc agtaccttat cgcgaagccg gaagaacctc ttaacctcta
     2221 gatgaaaaag cctgaactca ccgcgacgtc tgtcgagaag ttcctgatcg aaaagttcga
     2281 cagcgtctcc gacctgatgc agctctcgga gggcgaagaa tctcgtgctt tcagcttcga
     2341 tgtaggaggg cgtggatatg tcctgcgggt aaatagctgc gccgatggtt tctacaaaga
     2401 tcgttatgtt tatcggcact ttgcatcggc cgcgctcccg attccggaag tgcttgacat
     2461 tggggaattc agcgagagcc tgacctattg catctcccgc cgtgcacagg gtgtcacgtt
     2521 gcaagacctg cctgaaaccg aactgcccgc tgttctgcag ccggtcgcgg aggccatgga
     2581 tgcgatcgct gcggccgatc tcagccagac gagcgggttc ggcccattcg gaccgcaagg
     2641 aatcggtcaa tacactacat ggcgtgattt catatgcgcg attgctgatc cccatgtgta
     2701 tcactggcaa actgtgatgg acgacaccgt cagtgcgtcc gtcgcgcagg ctctcgatga
     2761 gctgatgctt tgggccgagg actgccccga agtccggcac ctcgtgcacg cggatttcgg
     2821 ctccaacaat gtcctgacgg acaatggccg cataacagcg gtcattgact ggagcgaggc
     2881 gatgttcggg gattcccaat acgaggtcgc caacatcttc ttctggaggc cgtggttggc
     2941 ttgtatggag cagcagacgc gctacttcga gcggaggcac ccggagcttg caggatcgcc
     3001 gcggctccgg gcgtatatgc tccgcattgg tcttgaccaa ctctatcaga gcttggttga
     3061 cggcaatttc gatgatgcag cttgggcgca gggtcgatgc gacgcaatcg tccgatccgg
     3121 agccgggact gtcgggcgta cacaaatcgc ccgcagaagc gcggccgtct ggaccgatgg
     3181 ctgtgtagaa gtactcgccg atagtggaaa ccgacgcccc agcactcgtc cgagggcaaa
     3241 ggaatagatg catggctttc gtgaccgggc ttcaaacaat gatgtgcgat ggtgtggttc
     3301 ccggttggcg gagtctttgt ctactttggt tgtctgtcgc aggtcggtag accgcaaatg
     3361 agcaactgat ggattgttgc cagcgatact ataattcaca tggatggtct ttgtcgatca
     3421 gtagctagtg agagagagag aacatctatc cacaatgtcg agtgtctatt agacatactc
     3481 cgagaataaa gtcaactgtg tctgtgatct aaagatcgat tcggcagtcg agtagcgtat
     3541 aacaactccg agtaccagcg aaagcacgtc gtgacaggag cagggctttg ccaactgcgc
     3601 aaccttgctt gaatgaggat acacggggtg caacatggct gtactgatcc atcgcaacca
     3661 aaatttctgt ttatagatca agctggtaga ttccaattac tccacctctt gcgcttctcc
     3721 atgacatgta agtgcacgtg gaaaccatac ccaaattgcc tacagctgcg gagcatgagc
     3781 ctatggcgat cagtctggtc atgttaacca gcctgtgctc tgacgttaat gcagaatagc
     3841 ttcgcgtgat cttacggcat tatacgtatg atcggtccac gatcagctag attatctagt
     3901 cagcttgatg tcatagctgt ttcctgaggc tcaatactga ccatttaaat catacctgac
     3961 ctccatagca gaaagtcaaa agcctccgac cggaggcttt tgacttgatc ggcacgtaag
     4021 aggttccaac tttcaccata atgaaataag atcactaccg ggcgtatttt ttgagttatc
     4081 gagattttca ggagctaagg aagctaaaat gagccatatt caacgggaaa cgtcttgctt
     4141 gaagccgcga ttaaattcca acatggatgc tgatttatat gggtataaat gggctcgcga
     4201 taatgtcggg caatcaggtg cgacaatcta tcgattgtat gggaagcccg atgcgccaga
     4261 gttgtttctg aaacatggca aaggtagcgt tgccaatgat gttacagatg agatggtcag
     4321 gctaaactgg ctgacggaat ttatgcctct tccgaccatc aagcatttta tccgtactcc
     4381 tgatgatgca tggttactca ccactgcgat cccagggaaa acagcattcc aggtattaga
     4441 agaatatcct gattcaggtg aaaatattgt tgatgcgctg gcagtgttcc tgcgccggtt
     4501 gcattcgatt cctgtttgta attgtccttt taacggcgat cgcgtatttc gtctcgctca
     4561 ggcgcaatca cgaatgaata acggtttggt tggtgcgagt gattttgatg acgagcgtaa
     4621 tggctggcct gttgaacaag tctggaaaga aatgcataaa ctcttgccat tctcaccgga
     4681 ttcagtcgtc actcatggtg atttctcact tgataacctt atttttgacg aggggaaatt
     4741 aataggttgt attgatgttg gacgagtcgg aatcgcagac cgataccagg atcttgccat
     4801 cctatggaac tgcctcggtg agttttctcc ttcattacag aaacggcttt ttcaaaaata
     4861 tggtattgat aatcctgata tgaataaatt gcagtttcac ttgatgctcg atgagttttt
     4921 ctaatgagga cctaaatgta atcacctggc tcaccttcgg gtgggccttt ctgcgttgct
     4981 ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgat gctcaagtca
     5041 gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct
     5101 cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc
     5161 gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt
     5221 tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc
     5281 cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc
     5341 cactggtaac
//
