LOCUS       sequence-226712-        3775 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Complete TU for hygromicin resistance in fungi, domesticated into 
            pUPD2 with AATG/GCTT barcodes. Used for gene KO with dual selection,
            according to FungalBraid modular DNA assembly for ATMT.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3775)
  AUTHORS   Hernanz-Koers M, Gandia M, Garrigues S, Manzanares P, Yenush L, 
            Orzaez D, Marcos JF
  TITLE     FungalBraid: A GoldenBraid-based modular cloning platform for the 
            assembly and exchange of DNA elements tailored to fungal synthetic 
            biology.
  JOURNAL   Fungal Genet Biol. 2018 Jul;116:51-61. doi: 
            10.1016/j.fgb.2018.04.010. Epub 2018 Apr 20.
  PUBMED    29680684
REFERENCE   2  (bases 1 to 3775)
  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: "Fungal 
            Genet Biol."; date: "2018-07"; volume: "116:51-61. doi"; pages: " 
            10.1016/j.fgb.2018.04.010. Epub 2018 Apr 20"
COMMENT     Sequence Label: FB012
FEATURES             Location/Qualifiers
     source          1..3775
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             367..1392
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     CDS             367..1392
                     /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"
     terminator      1712..1769
                     /label=his operon terminator
                     /note="This putative transcriptin terminator from the E. 
                     coli his operon has a 2-bp deletion introduced during 
                     synthesis. Its efficiency has not been determined."
     primer_bind     complement(1793..1810)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1964..2552)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2044..2063)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      complement(2735..2829)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(2850..3509)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     3424..3443
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     primer_bind     3671..3689
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     terminator      complement(3693..3736)
                     /label=bacterial terminator
                     /note="putative bacterial transcription terminator"
     misc_feature    3739..3760
                     /label=BioBrick prefix
                     /note="BioBrick prefix for parts that do not start with 
                     ""ATG"""
ORIGIN
        1 aatgaactga tattgaagga gcattttttg ggcttggctg gagctagtgg aggtcaacaa
       61 tgaatgccta ttttggttta gtcgtccagg cggtgagcac aaaatttgtg tcgtttgaca
      121 agatggttca tttaggcaac tggtcagatc agccccactt gtagcagtag cggcggcgct
      181 cgaagtgtga ctcttattag cagacaggaa cgaggacatt attatcatct gctgcttggt
      241 gcacgataac ttggtgcgtt tgtcaagcaa ggtaagtgga cgacccggtc ataccttctt
      301 aagttcgccc ttcctccctt tatttcagat tcaatctgac ttacctattc tacccaagca
      361 tccaaaatga aaaagcctga actcaccgcg acgtctgtcg agaagtttct gatcgaaaag
      421 ttcgacagcg tgtccgacct gatgcagctc tcggagggcg aagaatctcg tgctttcagc
      481 ttcgatgtag gagggcgtgg atatgtcctg cgggtaaata gctgcgccga tggtttctac
      541 aaagatcgtt atgtttatcg gcactttgca tcggccgcgc tcccgattcc ggaagtgctt
      601 gacattgggg aattcagcga gagcctgacc tattgcatct cccgccgtgc acagggtgtc
      661 acgttgcaag acctgcctga aaccgaactg cccgctgttc tgcagccggt cgcggaggcc
      721 atggatgcga tcgctgcggc cgatcttagc cagacgagcg ggttcggccc attcggaccg
      781 caaggaatcg gtcaatacac tacatggcgt gatttcatat gcgcgattgc tgatccccat
      841 gtgtatcact ggcaaactgt gatggacgac accgtcagtg cgtccgtcgc gcaggctctc
      901 gatgagctga tgctttgggc cgaggactgc cccgaagtcc ggcacctcgt gcacgcggat
      961 ttcggctcca acaatgtcct gacggacaat ggccgcataa cagcggtcat tgactggagc
     1021 gaggcaatgt tcggggattc ccaatacgag gtcgccaaca tcttcttctg gaggccgtgg
     1081 ttggcttgta tggagcagca gacgcgctac ttcgagcgga ggcatccgga gcttgcagga
     1141 tcgccacgac tccgggcgta tatgctccgc attggtcttg accaactcta tcagagcttg
     1201 gttgacggca atttcgatga tgcagcttgg gcgcagggtc gatgcgacgc aatcgtccga
     1261 tccggagccg ggactgtcgg gcgtacacaa atcgcccgca gaagcgcggc cgtctggacc
     1321 gatggctgtg tagaagtact cgccgatagt ggaaaccgac gccccagcac tcgtccgagg
     1381 gcaaaggaat gagcttcatc attccaccat cattccactc aacattcagg ctcctctgcg
     1441 cacgtaaagt gccaaaggca ataccctgct cggtggaatg ccgccgggct tgtcgatttt
     1501 acgcacatat gcgcattctt gacttgaagc ggaggagttc ttcgttgcgg gttacagtgt
     1561 tttaataaaa gaatggtcaa atcaaactgc tagatatacc tgtcagacac tctagttgtt
     1621 gacccctata ctcttaatac atcagacagt acatgcatgt tgcatgatga gctttgagac
     1681 cactcatcgc gactagtagc ggccgctgca gtccggcaaa aaagggcaag gtgtcaccac
     1741 cctgcccttt ttctttaaaa ccgaaaagat tacttcgcgt tatgcaggct tcctcgctca
     1801 ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg
     1861 taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc
     1921 agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccac aggctccgcc
     1981 cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac
     2041 tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc
     2101 tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata
     2161 gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc
     2221 acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca
     2281 acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag
     2341 cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta
     2401 gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg
     2461 gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc
     2521 agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt
     2581 ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa
     2641 ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat
     2701 atgagtaaac ttggtctgac agctcgaggc ttggattctc accaataaaa aacgcccggc
     2761 ggcaaccgag cgttctgaac aaatccagat ggagttctga ggtcattact ggatctatca
     2821 acaggagtcc aagcgagctc gatatcaaat tacgccccgc cctgccactc gtcgcagtac
     2881 tgttgtaatt cattaagcat tctgccgaca tggaagccat cacaaacggc atgatgaacc
     2941 tgaatcgcca gcggcatcag caccttgtcg ccttgcgtat aatatttgcc catggtgaaa
     3001 acgggggcga agaagttgtc catattggcc acgtttaaat caaaactggt gaaactcacc
     3061 cagggattgg ctgatacgaa aaacatattc tcaataaacc ctttagggaa ataggccagg
     3121 ttttcaccgt aacacgccac atcttgcgaa tatatgtgta gaaactgccg gaaatcgtcg
     3181 tggtattcac tccagagtga tgaaaacgtt tcagtttgct catggaaaac ggtgtaacaa
     3241 gggtgaacac tatcccatat caccagctca ccgtctttca ttgccatacg aaattccgga
     3301 tgagcattca tcaggcgggc aagaatgtga ataaaggccg gataaaactt gtgcttattt
     3361 ttctttacgg tctttaaaaa ggccgtaata tccagctgaa cggtctggtt ataggtacat
     3421 tgagcaactg actgaaatgc ctcaaaatgt tctttacgat gccattggga tatatcaacg
     3481 gtggtatatc cagtgatttt tttctccatt ttagcttcct tagctcctga aaatctcgat
     3541 aactcaaaaa atacgcccgg tagtgatctt atttcattat ggtgaaagtt ggaacctctt
     3601 acgtgcccga tcaactcgag tgccacctga cgtctaagaa accattatta tcatgacatt
     3661 aacctataaa aataggcgta tcacgaggca gaatttcaga taaaaaaaat ccttagcttt
     3721 cgctaaggat gatttctgga attcgcggcc gcttctagag cgatgagtgg tctcg
//
