LOCUS       40924_28227        8334 bp DNA     circular SYN 18-DEC-2018
DEFINITION  Cloning vector pLH331, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8334)
  AUTHORS   Xu Yongxue XY.
  TITLE     Development of a markerless genetic manipulation system in 
            aspergillus Niger and its application in generating high organic 
            acid producing strains
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 8334)
  AUTHORS   Xu Yongxue XY.
  TITLE     Direct Submission
  JOURNAL   Submitted (01-FEB-2018) College of Biotechnology, Tianjin University
REFERENCE   3  (bases 1 to 8334)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 8334)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (01-FEB-2018) College of Biotechnology, Tianjin University of 
            Science "
COMMENT     SGRef: number: 3; type: "Journal Article"
COMMENT     ##Assembly-Data-START##
            Sequencing Technology :: Sanger dideoxy sequencing 
            ##Assembly-Data-END##
FEATURES             Location/Qualifiers
     source          1..8334
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    6..39
                     /label=loxP
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        51..405
                     /label=trpC promoter
                     /note="promoter for Aspergillus nidulans trpC"
     CDS             410..1432
                     /codon_start=1
                     /label=HygR
                     /note="aminoglycoside phosphotransferase from E. coli"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANILFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     protein_bind    complement(1469..1502)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     primer_bind     complement(1542..1558)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_feature    1761..1785
                     /label=RB T-DNA repeat
                     /note="right border repeat from nopaline C58 T-DNA"
     CDS             3085..3711
                     /codon_start=1
                     /label=pVS1 StaA
                     /note="stability protein from the Pseudomonas plasmid pVS1
                     (Heeb et al., 2000)"
                     /translation="MKVIAVLNQKGGSGKTTIATHLARALQLAGADVLLVDSDPQGSAR
                     DWAAVREDQPLTVVGIDRPTIDRDVKAIGRRDFVVIDGAPQAADLAVSAIKAADFVLIP
                     VQPSPYDIWATADLVELVKQRIEVTDGRLQAAFVVSRAIKGTRIGGEVAEALAGYELPI
                     LESRITQRVSYPGTAAAGTTVLESEPEGDAAREVQALAAEIKSKLI"
     CDS             4148..5212
                     /codon_start=1
                     /label=pVS1 RepA
                     /note="replication protein from the Pseudomonas plasmid
                     pVS1 (Heeb et al., 2000)"
                     /translation="GRKPSGPVQIGAALGDDLVEKLKAAQAAQRQRIEAEARPGESWQA
                     AADRIRKESRQPPAAGAPSIRKPPKGDEQPDFFVPMLYDVGTRDSRSIMDVAVFRLSKR
                     DRRAGEVIRYELPDGHVEVSAGPAGMASVWDYDLVLMAVSHLTESMNRYREGKGDKPGR
                     VFRPHVADVLKFCRRADGGKQKDDLVETCIRLNTTHVAMQRTKKAKNGRLVTVSEGEAL
                     ISRYKIVKSETGRPEYIEIELADWMYREITEGKNPDVLTVHPDYFLIDPGIGRFLYRLA
                     RRAAGKAEARWLFKTIYERSGSAGEFKKFCFTVRKLIGSNDLPEYDLKEEAGQAGPILV
                     MRYRNLIEGEASAGS"
     rep_origin      5281..5475
                     /label=pVS1 oriV
                     /note="origin of replication for the Pseudomonas plasmid
                     pVS1 (Heeb et al., 2000)"
     misc_feature    5819..5959
                     /label=bom
                     /note="basis of mobility region from pBR322"
     rep_origin      complement(6145..6733)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(6823..7614)
                     /codon_start=1
                     /label=KanR
                     /note="aminoglycoside phosphotransferase"
                     /translation="MAKMRISPELKKLIEKYRCVKDTEGMSPAKVYKLVGENENLYLKM
                     TDSRYKGTTYDVEREKDMMLWLEGKLPVPKVLHFERHDGWSNLLMSEADGVLCSEEYED
                     EQSPEKIIELYAECIRLFHSIDISDCPYTNSLDSRLAELDYLLNNDLADVDCENWEEDT
                     PFKDPRELYDFLKTEKPEEELVFSHGDLGDSNIFVKDGKVSGFIDLGRSGRADKWYDIA
                     FCVRSIREDIGEEQYVELFFDLLGIKPDWEKIKYYILLDELF"
     misc_feature    8039..8063
                     /label=LB T-DNA repeat
                     /note="left border repeat from nopaline C58 T-DNA"
     polyA_signal    complement(8141..8315)
                     /label=CaMV poly(A) signal
                     /note="cauliflower mosaic virus polyadenylation signal"
ORIGIN
        1 gatccataac ttcgtataat gtatgctata cgaagttatg tcgacgttaa ctgatattga
       61 aggagcattt tttgggcttg gctggagcta gtggaggtca acaatgaatg cctattttgg
      121 tttagtcgtc caggcggtga gcacaaaatt tgtgtcgttt gacaagatgg ttcatttagg
      181 caactggtca gatcagcccc acttgtagca gtagcggcgg cgctcgaagt gtgactctta
      241 ttagcagaca ggaacgagga cattattatc atctgctgct tggtgcacga taacttggtg
      301 cgtttgtcaa gcaaggtaag tggacgaccc ggtcatacct tcttaagttc gcccttcctc
      361 cctttatttc agattcaatc tgacttacct attctaccca agcatccaaa tgaaaaagcc
      421 tgaactcacc gcgacgtctg tcgagaagtt tctgatcgaa aagttcgaca gcgtctccga
      481 cctgatgcag ctctcggagg gcgaagaatc tcgtgctttc agcttcgatg taggagggcg
      541 tggatatgtc ctgcgggtaa atagctgcgc cgatggtttc tacaaagatc gttatgttta
      601 tcggcacttt gcatcggccg cgctcccgat tccggaagtg cttgacattg gggagttcag
      661 cgagagcctg acctattgca tctcccgccg tgcacagggt gtcacgttgc aagacctgcc
      721 tgaaaccgaa ctgcccgctg ttctccagcc ggtcgcggag gccatggatg cgatcgctgc
      781 ggccgatctt agccagacga gcgggttcgg cccattcgga ccgcaaggaa tcggtcaata
      841 cactacatgg cgtgatttca tatgcgcgat tgctgatccc catgtgtatc actggcaaac
      901 tgtgatggac gacaccgtca gtgcgtccgt cgcgcaggct ctcgatgagc tgatgctttg
      961 ggccgaggac tgccccgaag tccggcacct cgtgcatgcg gatttcggct ccaacaatgt
     1021 cctgacggac aatggccgca taacagcggt cattgactgg agcgaggcga tgttcgggga
     1081 ttcccaatac gaggtcgcca acatcctctt ctggaggccg tggttggctt gtatggagca
     1141 gcagacgcgc tacttcgagc ggaggcatcc ggagcttgca ggatcgccgc gcctccgggc
     1201 gtatatgctc cgcattggtc ttgaccaact ctatcagagc ttggttgacg gcaatttcga
     1261 tgatgcagct tgggcgcagg gtcgatgcga cgcaatcgtc cgatccggag ccgggactgt
     1321 cgggcgtaca caaatcgccc gcagaagcgc ggccgtctgg accgatggct gtgtagaagt
     1381 actcgccgat agtggaaacc gacgccccag cactcgtccg agggcaaagg aatagagtag
     1441 atgccgaccg ggaaccagtt aacgtcgaat aacttcgtat aatgtatgct atacgaagtt
     1501 attctagaac tagtgggccc ctgcaggcat gcaagcttgg cactggccgt cgttttacaa
     1561 cgtcgtgact gggaaaaccc tggcgttacc caacttaatc gccttgcagc acatccccct
     1621 ttcgccagct ggcgtaatag cgaagaggcc cgcaccgatc gcccttccca acagttgcgc
     1681 agcctgaatg gcgaatgcta gagcagcttg agcttggatc agattgtcgt ttcccgcctt
     1741 cagtttaaac tatcagtgtt tgacaggata tattggcggg taaacctaag agaaaagagc
     1801 gtttattaga ataacggata tttaaaaggg cgtgaaaagg tttatccgtt cgtccatttg
     1861 tatgtgcatg ccaaccacag ggttcccctc gggatcaaag tactttgatc caacccctcc
     1921 gctgctatag tgcagtcggc ttctgacgtt cagtgcagcc gtcttctgaa aacgacatgt
     1981 cgcacaagtc ctaagttacg cgacaggctg ccgccctgcc cttttcctgg cgttttcttg
     2041 tcgcgtgttt tagtcgcata aagtagaata cttgcgacta gaaccggaga cattacgcca
     2101 tgaacaagag cgccgccgct ggcctgctgg gctatgcccg cgtcagcacc gacgaccagg
     2161 acttgaccaa ccaacgggcc gaactgcacg cggccggctg caccaagctg ttttccgaga
     2221 agatcaccgg caccaggcgc gaccgcccgg agctggccag gatgcttgac cacctacgcc
     2281 ctggcgacgt tgtgacagtg accaggctag accgcctggc ccgcagcacc cgcgacctac
     2341 tggacattgc cgagcgcatc caggaggccg gcgcgggcct gcgtagcctg gcagagccgt
     2401 gggccgacac caccacgccg gccggccgca tggtgttgac cgtgttcgcc ggcattgccg
     2461 agttcgagcg ttccctaatc atcgaccgca cccggagcgg gcgcgaggcc gccaaggccc
     2521 gaggcgtgaa gtttggcccc cgccctaccc tcaccccggc acagatcgcg cacgcccgcg
     2581 agctgatcga ccaggaaggc cgcaccgtga aagaggcggc tgcactgctt ggcgtgcatc
     2641 gctcgaccct gtaccgcgca cttgagcgca gcgaggaagt gacgcccacc gaggccaggc
     2701 ggcgcggtgc cttccgtgag gacgcattga ccgaggccga cgccctggcg gccgccgaga
     2761 atgaacgcca agaggaacaa gcatgaaacc gcaccaggac ggccaggacg aaccgttttt
     2821 cattaccgaa gagatcgagg cggagatgat cgcggccggg tacgtgttcg agccgcccgc
     2881 gcacgtctca accgtgcggc tgcatgaaat cctggccggt ttgtctgatg ccaagctggc
     2941 ggcctggccg gccagcttgg ccgctgaaga aaccgagcgc cgccgtctaa aaaggtgatg
     3001 tgtatttgag taaaacagct tgcgtcatgc ggtcgctgcg tatatgatgc gatgagtaaa
     3061 taaacaaata cgcaagggga acgcatgaag gttatcgctg tacttaacca gaaaggcggg
     3121 tcaggcaaga cgaccatcgc aacccatcta gcccgcgccc tgcaactcgc cggggccgat
     3181 gttctgttag tcgattccga tccccagggc agtgcccgcg attgggcggc cgtgcgggaa
     3241 gatcaaccgc taaccgttgt cggcatcgac cgcccgacga ttgaccgcga cgtgaaggcc
     3301 atcggccggc gcgacttcgt agtgatcgac ggagcgcccc aggcggcgga cttggctgtg
     3361 tccgcgatca aggcagccga cttcgtgctg attccggtgc agccaagccc ttacgacata
     3421 tgggccaccg ccgacctggt ggagctggtt aagcagcgca ttgaggtcac ggatggaagg
     3481 ctacaagcgg cctttgtcgt gtcgcgggcg atcaaaggca cgcgcatcgg cggtgaggtt
     3541 gccgaggcgc tggccgggta cgagctgccc attcttgagt cccgtatcac gcagcgcgtg
     3601 agctacccag gcactgccgc cgccggcaca accgttcttg aatcagaacc cgagggcgac
     3661 gctgcccgcg aggtccaggc gctggccgct gaaattaaat caaaactcat ttgagttaat
     3721 gaggtaaaga gaaaatgagc aaaagcacaa acacgctaag tgccggccgt ccgagcgcac
     3781 gcagcagcaa ggctgcaacg ttggccagcc tggcagacac gccagccatg aagcgggtca
     3841 actttcagtt gccggcggag gatcacacca agctgaagat gtacgcggta cgccaaggca
     3901 agaccattac cgagctgcta tctgaataca tcgcgcagct accagagtaa atgagcaaat
     3961 gaataaatga gtagatgaat tttagcggct aaaggaggcg gcatggaaaa tcaagaacaa
     4021 ccaggcaccg acgccgtgga atgccccatg tgtggaggaa cgggcggttg gccaggcgta
     4081 agcggctggg ttgtctgccg gccctgcaat ggcactggaa cccccaagcc cgaggaatcg
     4141 gcgtgacggt cgcaaaccat ccggcccggt acaaatcggc gcggcgctgg gtgatgacct
     4201 ggtggagaag ttgaaggccg cgcaggccgc ccagcggcaa cgcatcgagg cagaagcacg
     4261 ccccggtgaa tcgtggcaag cggccgctga tcgaatccgc aaagaatccc ggcaaccgcc
     4321 ggcagccggt gcgccgtcga ttaggaagcc gcccaagggc gacgagcaac cagatttttt
     4381 cgttccgatg ctctatgacg tgggcacccg cgatagtcgc agcatcatgg acgtggccgt
     4441 tttccgtctg tcgaagcgtg accgacgagc tggcgaggtg atccgctacg agcttccaga
     4501 cgggcacgta gaggtttccg cagggccggc cggcatggcc agtgtgtggg attacgacct
     4561 ggtactgatg gcggtttccc atctaaccga atccatgaac cgataccggg aagggaaggg
     4621 agacaagccc ggccgcgtgt tccgtccaca cgttgcggac gtactcaagt tctgccggcg
     4681 agccgatggc ggaaagcaga aagacgacct ggtagaaacc tgcattcggt taaacaccac
     4741 gcacgttgcc atgcagcgta cgaagaaggc caagaacggc cgcctggtga cggtatccga
     4801 gggtgaagcc ttgattagcc gctacaagat cgtaaagagc gaaaccgggc ggccggagta
     4861 catcgagatc gagctagctg attggatgta ccgcgagatc acagaaggca agaacccgga
     4921 cgtgctgacg gttcaccccg attacttttt gatcgatccc ggcatcggcc gttttctcta
     4981 ccgcctggca cgccgcgccg caggcaaggc agaagccaga tggttgttca agacgatcta
     5041 cgaacgcagt ggcagcgccg gagagttcaa gaagttctgt ttcaccgtgc gcaagctgat
     5101 cgggtcaaat gacctgccgg agtacgattt gaaggaggag gcggggcagg ctggcccgat
     5161 cctagtcatg cgctaccgca acctgatcga gggcgaagca tccgccggtt cctaatgtac
     5221 ggagcagatg ctagggcaaa ttgccctagc aggggaaaaa ggtcgaaaag gtctctttcc
     5281 tgtggatagc acgtacattg ggaacccaaa gccgtacatt gggaaccgga acccgtacat
     5341 tgggaaccca aagccgtaca ttgggaaccg gtcacacatg taagtgactg atataaaaga
     5401 gaaaaaaggc gatttttccg cctaaaactc tttaaaactt attaaaactc ttaaaacccg
     5461 cctggcctgt gcataactgt ctggccagcg cacagccgaa gagctgcaaa aagcgcctac
     5521 ccttcggtcg ctgcgctccc tacgccccgc cgcttcgcgt cggcctatcg cggccgctgg
     5581 ccgctcaaaa atggctggcc tacggccagg caatctacca gggcgcggac aagccgcgcc
     5641 gtcgccactc gaccgccggc gcccacatca aggcaccctg cctcgcgcgt ttcggtgatg
     5701 acggtgaaaa cctctgacac atgcagctcc cggagacggt cacagcttgt ctgtaagcgg
     5761 atgccgggag cagacaagcc cgtcagggcg cgtcagcggg tgttggcggg tgtcggggcg
     5821 cagccatgac ccagtcacgt agcgatagcg gagtgtatac tggcttaact atgcggcatc
     5881 agagcagatt gtactgagag tgcaccatat gcggtgtgaa ataccgcaca gatgcgtaag
     5941 gagaaaatac cgcatcaggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt
     6001 cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga
     6061 atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg
     6121 taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa
     6181 aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt
     6241 tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct
     6301 gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct
     6361 cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc
     6421 cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt
     6481 atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc
     6541 tacagagttc ttgaagtggt ggcctaacta cggctacact agaaggacag tatttggtat
     6601 ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa
     6661 acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa
     6721 aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga
     6781 aaactcacgt taagggattt tggtcatgca ttctaggtac taaaacaatt catccagtaa
     6841 aatataatat tttattttct cccaatcagg cttgatcccc agtaagtcaa aaaatagctc
     6901 gacatactgt tcttccccga tatcctccct gatcgaccgg acgcagaagg caatgtcata
     6961 ccacttgtcc gccctgccgc ttctcccaag atcaataaag ccacttactt tgccatcttt
     7021 cacaaagatg ttgctgtctc ccaggtcgcc gtgggaaaag acaagttcct cttcgggctt
     7081 ttccgtcttt aaaaaatcat acagctcgcg cggatcttta aatggagtgt cttcttccca
     7141 gttttcgcaa tccacatcgg ccagatcgtt attcagtaag taatccaatt cggctaagcg
     7201 gctgtctaag ctattcgtat agggacaatc cgatatgtcg atggagtgaa agagcctgat
     7261 gcactccgca tacagctcga taatcttttc agggctttgt tcatcttcat actcttccga
     7321 gcaaaggacg ccatcggcct cactcatgag cagattgctc cagccatcat gccgttcaaa
     7381 gtgcaggacc tttggaacag gcagctttcc ttccagccat agcatcatgt ccttttcccg
     7441 ttccacatca taggtggtcc ctttataccg gctgtccgtc atttttaaat ataggttttc
     7501 attttctccc accagcttat ataccttagc aggagacatt ccttccgtat cttttacgca
     7561 gcggtatttt tcgatcagtt ttttcaattc cggtgatatt ctcattttag ccatttatta
     7621 tttccttcct cttttctaca gtatttaaag ataccccaag aagctaatta taacaagacg
     7681 aactccaatt cactgttcct tgcattctaa aaccttaaat accagaaaac agctttttca
     7741 aagttgtttt caaagttggc gtataacata gtatcgacgg agccgatttt gaaaccgcgg
     7801 tgatcacagg cagcaacgct ctgtcatcgt tacaatcaac atgctaccct ccgcgagatc
     7861 atccgtgttt caaacccggc agcttagttg ccgttcttcc gaatagcatc ggtaacatga
     7921 gcaaagtctg ccgccttaca acggctctcc cgctgacgcc gtcccggact gatgggctgc
     7981 ctgtatcgag tggtgatttt gtgccgagct gccggtcggg gagctgttgg ctggctggtg
     8041 gcaggatata ttgtggtgta aacaaattga cgcttagaca acttaataac acattgcgga
     8101 cgtttttaat gtactgaatt aacgccgaat taattcgggg gatctggatt ttagtactgg
     8161 attttggttt taggaattag aaattttatt gatagaagta ttttacaaat acaaatacat
     8221 actaagggtt tcttatatgc tcaacacatg agcgaaaccc tataggaacc ctaattccct
     8281 tatctgggaa ctactcacac attattatgg agaaactcga ggaattcgag ctcg
//
