LOCUS       Exported                4833 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    pA-HYG OSCAR
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4833)
  AUTHORS   Paz Z, Garcia-Pedrajas MD, Andrews DL, Klosterman SJ, Baeza-Montanez
            L, Gold SE
  TITLE     One Step Construction of Agrobacterium-Recombination-ready-plasmids 
            (OSCAR), an efficient and robust tool for ATMT based gene deletion 
            construction in fungi.
  JOURNAL   Fungal Genet Biol. 2011 Mar 5. ():.
  PUBMED    21362493
REFERENCE   2  (bases 1 to 4833)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..4833
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     rep_origin      complement(3..458)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(140..161)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     352..371
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     585..607
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     599..616
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     600..616
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     626..645
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        626..644
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    complement(678..909)
                     /gene="mutant version of attP"
                     /label=attP4
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     916..932
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     916..932
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     CDS             complement(959..1984)
                     /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
                     GDSQYEVANILFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     protein_bind    complement(2361..2592)
                     /gene="mutant version of attP"
                     /label=attP1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction 
                     (pDONR(TM)201 version)"
     primer_bind     complement(2645..2665)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(2645..2663)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2684..2700)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2684..2700)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2697..2719)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2708..2724
                     /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)."
     promoter        complement(2732..2762)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2777..2798
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2915..2932)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3086..3674)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3166..3185)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(3845..4705)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     4468..4487
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4706..4810)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ctaaattgta agcgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc
       61 attttttaac caataggccg aaatcggcaa aatcccttat aaatcaaaag aatagaccga
      121 gatagggttg agtgttgttc cagtttggaa caagagtcca ctattaaaga acgtggactc
      181 caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg aaccatcacc
      241 ctaatcaagt tttttggggt cgaggtgccg taaagcacta aatcggaacc ctaaagggag
      301 cccccgattt agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa
      361 agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac
      421 cacacccgcc gcgcttaatg cgccgctaca gggcgcgtcc cattcgccat tcaggctgcg
      481 caactgttgg gaagggcgat cggtgcgggc ctcttcgcta ttacgccagc tggcgaaagg
      541 gggatgtgct gcaaggcgat taagttgggt aacgccaggg ttttcccagt cacgacgttg
      601 taaaacgacg gccagtgagc gcgcgtaata cgactcacta tagggcgaat tggagctcca
      661 ccgcggtgcc ggaattctac aggtcactaa taccatctaa gtagttgatt catagtgact
      721 gcatatgttg tgttttacag tattatgtag tctgtttttt atgcaaaatc taatttaata
      781 tattgatatt tatatcattt tacgtttctc gttcaacttt tctatacaaa gttggcatta
      841 taaaaaagca ttgcttatca atttgttgca acgaacaggt cactatcagt caaaataaaa
      901 tcattatttg gccgctctag aactagtgga tccaactggt tcccggtcgg catctactct
      961 attcctttgc cctcggacga gtgctggggc gtcggtttcc actatcggcg agtacttcta
     1021 cacagccatc ggtccagacg gccgcgcttc tgcgggcgat ttgtgtacgc ccgacagtcc
     1081 cggctccgga tcggacgatt gcgtcgcatc gaccctgcgc ccaagctgca tcatcgaaat
     1141 tgccgtcaac caagctctga tagagttggt caagaccaat gcggagcata tacgcccgga
     1201 ggcgcggcga tcctgcaagc tccggatgcc tccgctcgaa gtagcgcgtc tgctgctcca
     1261 tacaagccaa ccacggcctc cagaagagga tgttggcgac ctcgtattgg gaatccccga
     1321 acatcgcctc gctccagtca atgaccgctg ttatgcggcc attgtccgtc aggacattgt
     1381 tggagccgaa atccgcatgc acgaggtgcc ggacttcggg gcagtcctcg gcccaaagca
     1441 tcagctcatc gagagcctgc gcgacggacg cactgacggt gtcgtccatc acagtttgcc
     1501 agtgatacac atggggatca gcaatcgcgc atatgaaatc acgccatgta gtgtattgac
     1561 cgattccttg cggtccgaat gggccgaacc cgctcgtctg gctaagatcg gccgcagcga
     1621 tcgcatccat ggcctccgcg accggctgga gaacagcggg cagttcggtt tcaggcaggt
     1681 cttgcaacgt gacaccctgt gcacggcggg agatgcaata ggtcaggctc tcgctgaact
     1741 ccccaatgtc aagcacttcc ggaatcggga gcgcggccga tgcaaagtgc cgataaacat
     1801 aacgatcttt gtagaaacca tcggcgcagc tatttacccg caggacatat ccacgccctc
     1861 ctacatcgaa gctgaaagca cgagattctt cgccctccga gagctgcatc aggtcggaga
     1921 cgctgtcgaa cttttcgatc agaaacttct cgacagacgt cgcggtgagt tcaggctttt
     1981 tcatttggat gcttgggtag aataggtaag tcagattgaa tctgaaataa agggaggaag
     2041 ggcgaactta agaaggtatg accgggtcgt ccacttacct tgcttgacaa acgcaccaag
     2101 ttatcgtgca ccaagcagca gatgataata atgtcctcgt tcctgtctgc taataagagt
     2161 cacacttcga gcgccgccgc tactgctaca agtggggctg atctgaccag ttgcctaaat
     2221 gaaccatctt gtcaaacgac acaaattttg tgctcaccgc ctggacgact aaaccaaaat
     2281 aggcattcat tgttgacctc cactagctcc agccaagccc aaaaaatgct ccttcaatat
     2341 cagttgggct gcaggaattc tacaggtcac taataccatc taagtagttg attcatagtg
     2401 actgcatatg ttgtgtttta cagtattatg tagtctgttt tttatgcaaa atctaattta
     2461 atatattgat atttatatca ttttacgttt ctcgttcaac ttttttgtac aaagttggca
     2521 ttataaaaaa gcattgctta tcaatttgtt gcaacgaaca ggtcactatc agtcaaaata
     2581 aaatcattat ttggagctct agagcgtcga cctcgagggg gggcccggta cccagctttt
     2641 gttcccttta gtgagggtta attgcgcgct tggcgtaatc atggtcatag ctgtttcctg
     2701 tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagc ataaagtgta
     2761 aagcctgggg tgcctaatga gtgagctaac tcacattaat tgcgttgcgc tcactgcccg
     2821 ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga
     2881 gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg
     2941 tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag
     3001 aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc
     3061 gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca
     3121 aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt
     3181 ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc
     3241 tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc
     3301 tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc
     3361 ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact
     3421 tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg
     3481 ctacagagtt cttgaagtgg tggcctaact acggctacac tagaaggaca gtatttggta
     3541 tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca
     3601 aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa
     3661 aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg
     3721 aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc
     3781 ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg
     3841 acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat
     3901 ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg
     3961 gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa
     4021 taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca
     4081 tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc
     4141 gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt
     4201 cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa
     4261 aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat
     4321 cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct
     4381 tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga
     4441 gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag
     4501 tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga
     4561 gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca
     4621 ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg
     4681 cgacacggaa atgttgaata ctcatactct tcctttttca atattattga agcatttatc
     4741 agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag
     4801 gggttccgcg cacatttccc cgaaaagtgc cac
//
