LOCUS       sequence-181718-        5351 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  BB3 recipient for homologous integration in pyrG locus with linker 
            AG.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5351)
  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 5351)
  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: pMST639_BB3_L_AG_hyg_KanOri_pyrG5
FEATURES             Location/Qualifiers
     source          1..5351
                     /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             2223..3248
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     CDS             2223..3248
                     /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(3911..3927)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3911..3927)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     terminator      3975..4006
                     /label=tonB terminator
                     /note="bidirectional E. coli tonB-P14 transcription 
                     terminator"
     promoter        4007..4109
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     CDS             4110..4925
                     /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(4183..4202)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     terminator      4949..4976
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     rep_origin      complement(join(4988..5351,1..224))
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5068..5087)
                     /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 gtctcccgct 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 aaaatccaac cactgacggc tgccggctct
     1861 gccacccccc tccctccacc ccagaccacc tgcacactca gcgcgcagca tcacctaatc
     1921 ttggctcgcc ttcccgcagc tcaggttgtt ttttttttct ctctccctcg tcgaagccgc
     1981 ccttgttccc ttatttattt ccctctccat ccttgtctgc ctttggtcca tctgcccctt
     2041 tgtctgcatc tcttttgcac gcatcgcctt atcgtcgtct cttttttcac tcacgggagc
     2101 ttgacgatga cctgactcgt gagcctcacc tgctgatttc tctccccccc tcccgaccgg
     2161 cttgactttt gtttctcctc cagtacctta tcgcgaagcc ggaagaacct cttaacctct
     2221 agatgaaaaa gcctgaactc accgcgacgt ctgtcgagaa gttcctgatc gaaaagttcg
     2281 acagcgtctc cgacctgatg cagctctcgg agggcgaaga atctcgtgct ttcagcttcg
     2341 atgtaggagg gcgtggatat gtcctgcggg taaatagctg cgccgatggt ttctacaaag
     2401 atcgttatgt ttatcggcac tttgcatcgg ccgcgctccc gattccggaa gtgcttgaca
     2461 ttggggaatt cagcgagagc ctgacctatt gcatctcccg ccgtgcacag ggtgtcacgt
     2521 tgcaagacct gcctgaaacc gaactgcccg ctgttctgca gccggtcgcg gaggccatgg
     2581 atgcgatcgc tgcggccgat ctcagccaga cgagcgggtt cggcccattc ggaccgcaag
     2641 gaatcggtca atacactaca tggcgtgatt tcatatgcgc gattgctgat ccccatgtgt
     2701 atcactggca aactgtgatg gacgacaccg tcagtgcgtc cgtcgcgcag gctctcgatg
     2761 agctgatgct ttgggccgag gactgccccg aagtccggca cctcgtgcac gcggatttcg
     2821 gctccaacaa tgtcctgacg gacaatggcc gcataacagc ggtcattgac tggagcgagg
     2881 cgatgttcgg ggattcccaa tacgaggtcg ccaacatctt cttctggagg ccgtggttgg
     2941 cttgtatgga gcagcagacg cgctacttcg agcggaggca cccggagctt gcaggatcgc
     3001 cgcggctccg ggcgtatatg ctccgcattg gtcttgacca actctatcag agcttggttg
     3061 acggcaattt cgatgatgca gcttgggcgc agggtcgatg cgacgcaatc gtccgatccg
     3121 gagccgggac tgtcgggcgt acacaaatcg cccgcagaag cgcggccgtc tggaccgatg
     3181 gctgtgtaga agtactcgcc gatagtggaa accgacgccc cagcactcgt ccgagggcaa
     3241 aggaatagat gcatggcttt cgtgaccggg cttcaaacaa tgatgtgcga tggtgtggtt
     3301 cccggttggc ggagtctttg tctactttgg ttgtctgtcg caggtcggta gaccgcaaat
     3361 gagcaactga tggattgttg ccagcgatac tataattcac atggatggtc tttgtcgatc
     3421 agtagctagt gagagagaga gaacatctat ccacaatgtc gagtgtctat tagacatact
     3481 ccgagaataa agtcaactgt gtctgtgatc taaagatcga ttcggcagtc gagtagcgta
     3541 taacaactcc gagtaccagc gaaagcacgt cgtgacagga gcagggcttt gccaactgcg
     3601 caaccttgct tgaatgagga tacacggggt gcaacatggc tgtactgatc catcgcaacc
     3661 aaaatttctg tttatagatc aagctggtag attccaatta ctccacctct tgcgcttctc
     3721 catgacatgt aagtgcacgt ggaaaccata cccaaattgc ctacagctgc ggagcatgag
     3781 cctatggcga tcagtctggt catgttaacc agcctgtgct ctgacgttaa tgcagaatag
     3841 cttcgcgtga tcttacggca ttatacgtat gatcggtcca cgatcagcta gattatctag
     3901 tcagcttgat gtcatagctg tttcctgagg ctcaatactg accatttaaa tcatacctga
     3961 cctccatagc agaaagtcaa aagcctccga ccggaggctt ttgacttgat cggcacgtaa
     4021 gaggttccaa ctttcaccat aatgaaataa gatcactacc gggcgtattt tttgagttat
     4081 cgagattttc aggagctaag gaagctaaaa tgagccatat tcaacgggaa acgtcttgct
     4141 tgaagccgcg attaaattcc aacatggatg ctgatttata tgggtataaa tgggctcgcg
     4201 ataatgtcgg gcaatcaggt gcgacaatct atcgattgta tgggaagccc gatgcgccag
     4261 agttgtttct gaaacatggc aaaggtagcg ttgccaatga tgttacagat gagatggtca
     4321 ggctaaactg gctgacggaa tttatgcctc ttccgaccat caagcatttt atccgtactc
     4381 ctgatgatgc atggttactc accactgcga tcccagggaa aacagcattc caggtattag
     4441 aagaatatcc tgattcaggt gaaaatattg ttgatgcgct ggcagtgttc ctgcgccggt
     4501 tgcattcgat tcctgtttgt aattgtcctt ttaacggcga tcgcgtattt cgtctcgctc
     4561 aggcgcaatc acgaatgaat aacggtttgg ttggtgcgag tgattttgat gacgagcgta
     4621 atggctggcc tgttgaacaa gtctggaaag aaatgcataa actcttgcca ttctcaccgg
     4681 attcagtcgt cactcatggt gatttctcac ttgataacct tatttttgac gaggggaaat
     4741 taataggttg tattgatgtt ggacgagtcg gaatcgcaga ccgataccag gatcttgcca
     4801 tcctatggaa ctgcctcggt gagttttctc cttcattaca gaaacggctt tttcaaaaat
     4861 atggtattga taatcctgat atgaataaat tgcagtttca cttgatgctc gatgagtttt
     4921 tctaatgagg acctaaatgt aatcacctgg ctcaccttcg ggtgggcctt tctgcgttgc
     4981 tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga tgctcaagtc
     5041 agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc
     5101 tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt
     5161 cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg
     5221 ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat
     5281 ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag
     5341 ccactggtaa c
//
