LOCUS       sequence-177623-        6784 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Complements mutations in Aspergillus nidulans gene biA (AN6644) 
            (mutants require biotin).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6784)
  AUTHORS   Dohn JW Jr, Grubbs AW, Oakley CE, Oakley BR
  TITLE     New multi-marker strains and complementing genes for Aspergillus 
            nidulans molecular biology.
  JOURNAL   Fungal Genet Biol. 2018 Feb;111:1-6. doi: 10.1016/j.fgb.2018.01.003.
            Epub 2018 Jan 5.
  PUBMED    29309843
REFERENCE   2  (bases 1 to 6784)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Fungal 
            Genet Biol."; date: "2018-02"; volume: "111:1-6. doi"; pages: " 
            10.1016/j.fgb.2018.01.003. Epub 2018 Jan 5"
COMMENT     Sequence Label: pLO101
FEATURES             Location/Qualifiers
     source          1..6784
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    1..22
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(37..42,43..60,61..67)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    75..91
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     80..102
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     99..115
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     99..115
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        133..151
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     133..150
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     CDS             3493..3504
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(3566..3585)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3567..3585)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3592..3609)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3592..3608)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3601..3623)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     CDS             3746..4048
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="QFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDKV
                     SRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     3927..3946
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             4397..5191
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             4397..5191
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4451..4470)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5061..5080
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     regulatory      5391..5400
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             5397..5771
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     rep_origin      5909..6497
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6398..6417
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6651..6668
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 taatgtgagt tagctcactc attaggcacc ccaggcttta cactttatgc ttccggctcg
       61 tatgttgtgt ggaattgtga gcggataaca atttcacaca ggaaacagct atgaccatga
      121 ttacgccaag ctatttaggt gacactatag aatactcaag ctatgcatca agcttggtac
      181 cgagctcgga tccactagta acggccgcca gtgtgctgga attcgccctt ctgtctgaga
      241 ggaggcacta cagtctcttg acaggcggtt ctttgctccg gaatactcca tagatgtgca
      301 tggaccatag ttcaccacta ctagacaaat cttactctgt acaatgactc actgggccgc
      361 tactccgtac actaccctac caagcccgcc cgcagccgct cctcaatcac ccgcagtgcc
      421 tcgggcctgg aggtcacact agacattagg tagagcacgt ttccgagcac ccgggaatga
      481 acgttgaagc ctgggccacc cgtcagaagg atctcttgga gacccttggc cgccgtcgaa
      541 gtgtatcctg tgggtcatca catgttagtc gtcacatcgt ttcagggcag ggaagagaga
      601 tgagtaggaa agagaagaaa agtgagaaag gtagacgtac caccgccatg cgcatccttc
      661 aacgcaatca ccaacaccgt cccaatggca aacaccccct ccacccccgg cgcaaacgac
      721 agatccttca ccagcccctg cgaccacaca ctccagacct caggggcatc cccgcagcgg
      781 gaggcctcgc tggccatggc cgcagtcgac tgccaatccc gacggtaggc gtcccaggcc
      841 ccacggcgct ccatctgcat catcgtccgg agcgaatcca ccgccacctg gcagcccacc
      901 gcatgggcgg tgtagctatg gccatgcagg agcgcgtcgc tcttttccgg gctggaaaac
      961 gccgcgaaga tctcgttgct ggccatggtc gtgcacagcg gcaccaggcc cccggtgagc
     1021 agcttggcgt tgaccacaat gtcgctgtcg atgttcagga acgacgcgga ggatcggcgc
     1081 ccgagccggt agagacctgt aaagacctcg tccaggatca ccgggaggcc agaccaggag
     1141 tcttctccgc ggggggcggc ggcggcgtta gcggcgaaga gttctggatg cccgcggacg
     1201 acgtcggcca gacagcgctg gaagaggggg tcactgatcc gttagtatgc gccagtcatg
     1261 gacgggataa tatggatagg ctggtacgaa ccagaagagc atgcccccag cgcccagaat
     1321 gacgggctcc ataatgagcg atccgaattt catcccctgc tcgcgcacca ggccctcgat
     1381 cgtgcgtcgg atgtacttgt gatagcgttt cccggcgtcc gactgcaccc gctcctccac
     1441 atcaaagacg ctgctcagcg tagggaattc caggtccgcg cctaatgcgc cgtccacctc
     1501 gggtggcatc tggatcttcc acaccccctg cgccatcttg acctgcggga aatcaaacca
     1561 gtacccgcgc ccgcggtacc actcgacctt tttgttgtac gtcgacggct ctgagcagtc
     1621 catcacccca atggtatccc catggtagct gcccttgagc ccgaggatgc tgatctgctc
     1681 ggcgctggcg tcccatccgt agcgcgcaca cgccgcccgc aggcccatct tcaccgcgac
     1741 ctccatgccc gtactcccat tgtccgtgta gaacacctta cccagacgcg ggttgcccat
     1801 ctcattgagc agcgtctccg cgagcgtcag cgccggctcg tggatgttcc cggggaacat
     1861 gacgtgcccg tagcggccgg ccgcgtaggc cgcggacagc gccagctcgg ggttgccatg
     1921 gcccaggccc tgcgtccacc aggacgccga cccatcaaac gtcgcctgca gcttgccccc
     1981 cgcgtaggtc tggaagtagt cgtcgtaggc cgagtcgatc ggcgtgatgt ccttgggcgc
     2041 catgccgtgg tgctgcgtga acgggtacca gatggcatcg tgggcgcgac cggccatcgc
     2101 ctcgagacgc tcgatgcggc tcgcgttctt cgtcgccagc tcctccagca gtccggctgt
     2161 gtcggtgccc tgcgcgacgc ggtggtagta ctcagccata gcctcggcat cgcgcgcggc
     2221 ggcgtctgga tcttggaccg ccggtcgcgc gggcggcgca ggtagcggca gcaacgggat
     2281 gctctttttc tggaagtagt cgcgcagata ctcgtggttt ttgtagtagt cgtcgcggaa
     2341 gagcaggaca gagtggacgt cgtagccccg cagcaacagc gactcgtacg cggagatgga
     2401 cgaggaaatc ccgcccaggc gagagtcggc gaccagcacg attggcagcc gcagcggccg
     2461 gtagagatct gcctgggagt ttccatttgg tccgggcgaa tgcacgccgc cggcggtctc
     2521 taccagggcg aagttgacgc cgttgctcgc ccagtcggag agggtcgagt ggacggacgt
     2581 gatgatatca tcgtcggggg gtacctttgt tgtcagcact tgatagcccg gggactgtaa
     2641 atggcagagg atttacagag aagccttgca cggcgagatg cgggctgacg ggctcgttga
     2701 attgatatag acatttggtg atggtgcctg cggcgtagcg ctggatgtgg ctgtcgaggg
     2761 tgagtcaatg gatccaagag aagaacagca tacataccga tcatccgcat cgtctagagg
     2821 gccggttgag actggtttca ggaatgcagc ctgtccctgg gtcttctggc gctggacggc
     2881 gttgcagagc accgtcgaga ctatagtctt gccgacatcg gtgttggcgc cgtagacctg
     2941 gtgggcacgc agcgagcgcc acagagcagc tcccactggc ggcatggcaa tattgaaaga
     3001 ttagcaatgg aaggagaaac aggaaggact acagaactac atggagagaa tgagtcgatg
     3061 gagtgggaat gacgccgaag aagagccgcg gtaggctgag ggtgcctgag gcagcaacgg
     3121 ccaccagcac aaccagcaac aagacagaaa taactataca gctctcttca gtttacttga
     3181 gcatcaattg ccacctgaaa gaattggatt gttaagcatt acctcaggtt tatccacttg
     3241 taaggccgcc aggcgtcccg gctaggagta tgtactccgt tcacctggag gcctgacaca
     3301 gtcatacacg ttggatgaca ttcgcctccg tacaacatcc aacacgacat aaggcaacca
     3361 attcttcatc atccgttgaa ttagtcaatt gctctcatat gttggtcatg aggatcagtt
     3421 actccgtact ccgtctcccc gacttgttga acccagcggt tcctgactgg gtccttgaag
     3481 agggtgaaga gcattgaagg gcgaattctg cagatatcca tcacactggc ggccgctcga
     3541 gcatgcatct agagggccca attcgcccta tagtgagtcg tattacaatt cactggccgt
     3601 cgttttacaa cgtcgtgact gggaaaaccc tggcgttacc caacttaatc gccttgcagc
     3661 acatccccct ttcgccagct ggcgtaatag cgaagaggcc cgcaccgatc gcccttccca
     3721 acagttgcgc agcctatacg tacggcagtt taaggtttac acctataaaa gagagagccg
     3781 ttatcgtctg tttgtggatg tacagagtga tattattgac acgccggggc gacggatggt
     3841 gatccccctg gccagtgcac gtctgctgtc agataaagtc tcccgtgaac tttacccggt
     3901 ggtgcatatc ggggatgaaa gctggcgcat gatgaccacc gatatggcca gtgtgccggt
     3961 ctccgttatc ggggaagaag tggctgatct cagccaccgc gaaaatgaca tcaaaaacgc
     4021 cattaacctg atgttctggg gaatataaat gtcaggcatg agattatcaa aaaggatctt
     4081 cacctagatc cttttcacgt agaaagccag tccgcagaaa cggtgctgac cccggatgaa
     4141 tgtcagctac tgggctatct ggacaaggga aaacgcaagc gcaaagagaa agcaggtagc
     4201 ttgcagtggg cttacatggc gatagctaga ctgggcggtt ttatggacag caagcgaacc
     4261 ggaattgcca gctggggcgc cctctggtaa ggttgggaag ccctgcaaag taaactggat
     4321 ggctttcttg ccgccaagga tctgatggcg caggggatca agctctgatc aagagacagg
     4381 atgaggatcg tttcgcatga ttgaacaaga tggattgcac gcaggttctc cggccgcttg
     4441 ggtggagagg ctattcggct atgactgggc acaacagaca atcggctgct ctgatgccgc
     4501 cgtgttccgg ctgtcagcgc aggggcgccc ggttcttttt gtcaagaccg acctgtccgg
     4561 tgccctgaat gaactgcaag acgaggcagc gcggctatcg tggctggcca cgacgggcgt
     4621 tccttgcgca gctgtgctcg acgttgtcac tgaagcggga agggactggc tgctattggg
     4681 cgaagtgccg gggcaggatc tcctgtcatc tcaccttgct cctgccgaga aagtatccat
     4741 catggctgat gcaatgcggc ggctgcatac gcttgatccg gctacctgcc cattcgacca
     4801 ccaagcgaaa catcgcatcg agcgagcacg tactcggatg gaagccggtc ttgtcgatca
     4861 ggatgatctg gacgaagagc atcaggggct cgcgccagcc gaactgttcg ccaggctcaa
     4921 ggcgagcatg cccgacggcg aggatctcgt cgtgacccat ggcgatgcct gcttgccgaa
     4981 tatcatggtg gaaaatggcc gcttttctgg attcatcgac tgtggccggc tgggtgtggc
     5041 ggaccgctat caggacatag cgttggctac ccgtgatatt gctgaagagc ttggcggcga
     5101 atgggctgac cgcttcctcg tgctttacgg tatcgccgct cccgattcgc agcgcatcgc
     5161 cttctatcgc cttcttgacg agttcttctg aattattaac gcttacaatt tcctgatgcg
     5221 gtattttctc cttacgcatc tgtgcggtat ttcacaccgc atcaggtggc acttttcggg
     5281 gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc
     5341 tcatgagaca ataaccctga taaatgcttc aataatagca cgtgaggagg gccaccatgg
     5401 ccaagttgac cagtgccgtt ccggtgctca ccgcgcgcga cgtcgccgga gcggtcgagt
     5461 tctggaccga ccggctcggg ttctcccggg acttcgtgga ggacgacttc gccggtgtgg
     5521 tccgggacga cgtgaccctg ttcatcagcg cggtccagga ccaggtggtg ccggacaaca
     5581 ccctggcctg ggtgtgggtg cgcggcctgg acgagctgta cgccgagtgg tcggaggtcg
     5641 tgtccacgaa cttccgggac gcctccgggc cggccatgac cgagatcggc gagcagccgt
     5701 gggggcggga gttcgccctg cgcgacccgg ccggcaactg cgtgcacttc gtggccgagg
     5761 agcaggactg acacgtgcta aaacttcatt tttaatttaa aaggatctag gtgaagatcc
     5821 tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag
     5881 accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct
     5941 gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac
     6001 caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat actgttcttc
     6061 tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg
     6121 ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt
     6181 tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt
     6241 gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc
     6301 tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca
     6361 gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata
     6421 gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg
     6481 ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct
     6541 ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat aaccgtatta
     6601 ccgcctttga gtgagctgat accgctcgcc gcagccgaac gaccgagcgc agcgagtcag
     6661 tgagcgagga agcggaagag cgcccaatac gcaaaccgcc tctccccgcg cgttggccga
     6721 ttcattaatg cagctggcac gacaggtttc ccgactggaa agcgggcagt gagcgcaacg
     6781 caat
//
