LOCUS       sequence-237149-        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"
     terminator      complement(1..44)
                     /label=bacterial terminator
                     /note="putative bacterial transcription terminator"
     misc_feature    47..68
                     /label=BioBrick prefix
                     /note="BioBrick prefix for parts that do not start with 
                     ""ATG"""
     CDS             450..1475
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     CDS             450..1475
                     /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      1795..1852
                     /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(1876..1893)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2047..2635)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2127..2146)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      complement(2818..2912)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(2933..3592)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     3507..3526
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     primer_bind     3754..3772
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 atttcagata aaaaaaatcc ttagctttcg ctaaggatga tttctggaat tcgcggccgc
       61 ttctagagcg atgagtggtc tcgaatgaac tgatattgaa ggagcatttt ttgggcttgg
      121 ctggagctag tggaggtcaa caatgaatgc ctattttggt ttagtcgtcc aggcggtgag
      181 cacaaaattt gtgtcgtttg acaagatggt tcatttaggc aactggtcag atcagcccca
      241 cttgtagcag tagcggcggc gctcgaagtg tgactcttat tagcagacag gaacgaggac
      301 attattatca tctgctgctt ggtgcacgat aacttggtgc gtttgtcaag caaggtaagt
      361 ggacgacccg gtcatacctt cttaagttcg cccttcctcc ctttatttca gattcaatct
      421 gacttaccta ttctacccaa gcatccaaaa tgaaaaagcc tgaactcacc gcgacgtctg
      481 tcgagaagtt tctgatcgaa aagttcgaca gcgtgtccga cctgatgcag ctctcggagg
      541 gcgaagaatc tcgtgctttc agcttcgatg taggagggcg tggatatgtc ctgcgggtaa
      601 atagctgcgc cgatggtttc tacaaagatc gttatgttta tcggcacttt gcatcggccg
      661 cgctcccgat tccggaagtg cttgacattg gggaattcag cgagagcctg acctattgca
      721 tctcccgccg tgcacagggt gtcacgttgc aagacctgcc tgaaaccgaa ctgcccgctg
      781 ttctgcagcc ggtcgcggag gccatggatg cgatcgctgc ggccgatctt agccagacga
      841 gcgggttcgg cccattcgga ccgcaaggaa tcggtcaata cactacatgg cgtgatttca
      901 tatgcgcgat tgctgatccc catgtgtatc actggcaaac tgtgatggac gacaccgtca
      961 gtgcgtccgt cgcgcaggct ctcgatgagc tgatgctttg ggccgaggac tgccccgaag
     1021 tccggcacct cgtgcacgcg gatttcggct ccaacaatgt cctgacggac aatggccgca
     1081 taacagcggt cattgactgg agcgaggcaa tgttcgggga ttcccaatac gaggtcgcca
     1141 acatcttctt ctggaggccg tggttggctt gtatggagca gcagacgcgc tacttcgagc
     1201 ggaggcatcc ggagcttgca ggatcgccac gactccgggc gtatatgctc cgcattggtc
     1261 ttgaccaact ctatcagagc ttggttgacg gcaatttcga tgatgcagct tgggcgcagg
     1321 gtcgatgcga cgcaatcgtc cgatccggag ccgggactgt cgggcgtaca caaatcgccc
     1381 gcagaagcgc ggccgtctgg accgatggct gtgtagaagt actcgccgat agtggaaacc
     1441 gacgccccag cactcgtccg agggcaaagg aatgagcttc atcattccac catcattcca
     1501 ctcaacattc aggctcctct gcgcacgtaa agtgccaaag gcaataccct gctcggtgga
     1561 atgccgccgg gcttgtcgat tttacgcaca tatgcgcatt cttgacttga agcggaggag
     1621 ttcttcgttg cgggttacag tgttttaata aaagaatggt caaatcaaac tgctagatat
     1681 acctgtcaga cactctagtt gttgacccct atactcttaa tacatcagac agtacatgca
     1741 tgttgcatga tgagctttga gaccactcat cgcgactagt agcggccgct gcagtccggc
     1801 aaaaaagggc aaggtgtcac caccctgccc tttttcttta aaaccgaaaa gattacttcg
     1861 cgttatgcag gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc
     1921 ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg
     1981 aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct
     2041 ggcgtttttc cacaggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca
     2101 gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct
     2161 cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc
     2221 gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt
     2281 tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc
     2341 cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc
     2401 cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg
     2461 gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc
     2521 agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag
     2581 cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga
     2641 tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat
     2701 tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag
     2761 ttttaaatca atctaaagta tatatgagta aacttggtct gacagctcga ggcttggatt
     2821 ctcaccaata aaaaacgccc ggcggcaacc gagcgttctg aacaaatcca gatggagttc
     2881 tgaggtcatt actggatcta tcaacaggag tccaagcgag ctcgatatca aattacgccc
     2941 cgccctgcca ctcgtcgcag tactgttgta attcattaag cattctgccg acatggaagc
     3001 catcacaaac ggcatgatga acctgaatcg ccagcggcat cagcaccttg tcgccttgcg
     3061 tataatattt gcccatggtg aaaacggggg cgaagaagtt gtccatattg gccacgttta
     3121 aatcaaaact ggtgaaactc acccagggat tggctgatac gaaaaacata ttctcaataa
     3181 accctttagg gaaataggcc aggttttcac cgtaacacgc cacatcttgc gaatatatgt
     3241 gtagaaactg ccggaaatcg tcgtggtatt cactccagag tgatgaaaac gtttcagttt
     3301 gctcatggaa aacggtgtaa caagggtgaa cactatccca tatcaccagc tcaccgtctt
     3361 tcattgccat acgaaattcc ggatgagcat tcatcaggcg ggcaagaatg tgaataaagg
     3421 ccggataaaa cttgtgctta tttttcttta cggtctttaa aaaggccgta atatccagct
     3481 gaacggtctg gttataggta cattgagcaa ctgactgaaa tgcctcaaaa tgttctttac
     3541 gatgccattg ggatatatca acggtggtat atccagtgat ttttttctcc attttagctt
     3601 ccttagctcc tgaaaatctc gataactcaa aaaatacgcc cggtagtgat cttatttcat
     3661 tatggtgaaa gttggaacct cttacgtgcc cgatcaactc gagtgccacc tgacgtctaa
     3721 gaaaccatta ttatcatgac attaacctat aaaaataggc gtatcacgag gcaga
//
