LOCUS       Exported                7523 bp ds-DNA     circular SYN 12-SEP-2024
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7523)
  AUTHORS   M. Tominaga et al
  TITLE     Direct Submission
  JOURNAL   Exported Monday, Oct 7, 2024 from SnapGene 5.0.8
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7523
                     /organism="recombinant plasmid"
                     /lab_host="Pichia pastoris"
                     /mol_type="other DNA"
     promoter        96..200
                     /gene="bla"
                     /label=AmpR promoter
                     /note="color: #ffffff; direction: RIGHT"
     CDS             201..1061
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /note="This feature has 2 segments:
                     1:  201 ..  269 / #ccffcc / signal sequence
                     2:  270 .. 1061 / #ccffcc
                     Cleavage site after base 269"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      1232..1820
                     /direction=RIGHT
                     /label=ori (ColE1)
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
                     /note="color: #ffff00"
     protein_bind    2108..2129
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /note="color: #31849b"
     promoter        2144..2174
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
                     /note="This forward directional feature has 3 segments:
                     1: 2144 .. 2149 / #ffffff / -35
                     2: 2150 .. 2167 / #ffffff
                     3: 2168 .. 2174 / #ffffff / -10"
     protein_bind    2182..2198
                     /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)."
                     /note="color: #31849b"
     primer_bind     2206..2222
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
                     /note="color: #a020f0; direction: RIGHT"
     misc_feature    2241..2478
                     /label=KpARG4-UTR
                     /note="color: #a6acb3"
     CDS             2479..3876
                     /codon_start=1
                     /label=KpARG4
                     /note="color: #993366"
                     /translation="MSNQEEGLKLWGGRFTGATDPLMDLYNASLPYDKKMYKVDLEGTK
                     VYTEGLEKINLLTKDELSEIHRGLKLIEAEWAEGKFVEKPGDEDIHTANERRLGELIGR
                     GISGKVHTGRSRNDQVATDMRLYVRDNLTQLADYLKQFIQVIIKRAEQEIDVLMPGYTH
                     LQRAQPIRWSHWLSMYATYFTEDYERLNQIVKRLNKSPLGAGALAGHPYGIDREYIAER
                     LGFDSVIGNSLAAVSDRDFVVETMFWSSLFMNHISRFSEDLIIYSTGEFGFIKLADAYS
                     TGSSLMPQKKNPDSLELLRGKSGRCFGALAGFLMSIKSIPSTYNKDMQEDKEPLFDTLI
                     TVEHSILIASGVVSTLNIDAERMKNALTMDMLATDLADYLVRRGVPFRETHHISGECVR
                     QAEELNLSGIDQLSLEQLKSIDSRFEADVASTFDFEASVEKRTATGGTSKTAVLKQLDA
                     LNEKLES"
     misc_feature    join(3889..3952,3975..3994,4016..4035,4038..4057,4060..4079,
                     4131..4150,4173..4192,4214..4233,4236..4255,4258..4277,
                     4288..4453)
                     /label=KpHsl10.3
                     /note="This feature has 14 segments:
                     1: 3889 .. 3932 / #------
                     2: 3933 .. 3952 / #ffcc00 / luxO(original)
                     3: 3975 .. 3994 / #ffcc00 / luxO(original)
                     4: 4016 .. 4035 / #ffcc00 / luxO(original)
                     5: 4038 .. 4057 / #ffcc00 / luxO(original)
                     6: 4060 .. 4079 / #ffcc00 / luxO(original)
                     7: 4131 .. 4150 / #ffcc00 / luxO(original)
                     8: 4173 .. 4192 / #ffcc00 / luxO(original)
                     9: 4214 .. 4233 / #ff9900 / luxO(VF1161)
                     10: 4236 .. 4255 / #ff9900 / luxO(VF1161)
                     11: 4258 .. 4277 / #ff9900 / luxO(VF1161)
                     12: 4288 .. 4291 / #666699 / KpAOX1 promoter(-166_-163) 13:
                     4292 .. 4300 / #ff0000 / TATA box
                     14: 4301 .. 4453 / #666699 / KpAOX1 promoter(-153_-1)"
     primer_bind     3890..3916
                     /label=MT1475
                     /note="color: 黒; sequence: GCgTGTTTTTTATTCCTTAAGTtacgc;
                     added: 2024-08-31"
     primer_bind     complement(3906..3925)
                     /label=MT1474
                     /note="color: 黒; sequence: 
                     ttcgaccttgcgtaACTTAAcattatcatccggatcacataCG; added: 
                     2024-08-31"
     primer_bind     complement(3953..3967)
                     /label=MT1474
                     /note="color: 黒; sequence: 
                     ttcgaccttgcgtaACTTAAcattatcatccggatcacataCG; added: 
                     2024-08-31"
     primer_bind     complement(4081..4123)
                     /label=MT1474
                     /note="color: 黒; sequence: 
                     ttcgaccttgcgtaACTTAAcattatcatccggatcacataCG; added: 
                     2024-08-31"
     primer_bind     complement(4151..4165)
                     /label=MT1474
                     /note="color: 黒; sequence: 
                     ttcgaccttgcgtaACTTAAcattatcatccggatcacataCG; added: 
                     2024-08-31"
     CDS             4460..5176
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /note="This feature has 3 segments:
                     1: 4460 .. 4462 / #05fc14
                     2: 4463 .. 4465 / #05fc14 / 1a
                     3: 4466 .. 5176 / #05fc14"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     terminator      5254..5500
                     /gene="Pichia pastoris AOX1"
                     /label=AOX1 terminator
                     /note="transcription terminator for AOX1"
                     /note="color: #ffffff; direction: RIGHT"
     promoter        5547..5896
                     /label=ScTEF1
                     /note="color: #ffffff; direction: RIGHT"
     CDS             5897..6934
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /note="color: #ccffcc"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     EVGRV"
     primer_bind     complement(7129..7145)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
                     /note="color: #a020f0; direction: LEFT"
ORIGIN
        1 gacgaaaggg cctcgtgata cgcctatttt tataggttaa tgtcatgata ataatggttt
       61 cttagacgtc aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt
      121 tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat
      181 aatattgaaa aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt
      241 ttgcggcatt ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg
      301 ctgaagatca gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga
      361 tccttgagag ttttcgcccc gaagaacgtt ttccaatgat gagcactttt aaagttctgc
      421 tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga gcaactcggt cgccgcatac
      481 actattctca gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg
      541 gcatgacagt aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca
      601 acttacttct gacaacgatc ggaggaccga aggagctaac cgcttttttg cacaacatgg
      661 gggatcatgt aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg
      721 acgagcgtga caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg
      781 gcgaactact tactctagct tcccggcaac aattaataga ctggatggag gcggataaag
      841 ttgcaggacc acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg
      901 gagccggtga gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct
      961 cccgtatcgt agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac
     1021 agatcgctga gataggtgcc tcactgatta agcattggta actgtcagac caagtttact
     1081 catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga
     1141 tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt
     1201 cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct
     1261 gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc
     1321 taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgttc
     1381 ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc
     1441 tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg
     1501 ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt
     1561 cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg
     1621 agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg
     1681 gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt
     1741 atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag
     1801 gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt
     1861 gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg gataaccgta
     1921 ttaccgcctt tgagtgagct gataccgctc gccgcagccg aacgaccgag cgcagcgagt
     1981 cagtgagcga ggaagcggaa gagcgcccaa tacgcaaacc gcctctcccc gcgcgttggc
     2041 cgattcatta atgcagctgg cacgacaggt ttcccgactg gaaagcgggc agtgagcgca
     2101 acgcaattaa tgtgagttag ctcactcatt aggcacccca ggctttacac tttatgcttc
     2161 cggctcgtat gttgtgtgga attgtgagcg gataacaatt tcacacagga aacagctatg
     2221 accatgatta cgccgctagc catggttgtg cgacagattc actgtgaaag actgttcatt
     2281 atacccacgt ttcactggga gatgtaagcc ttaggtgttt taccctgatt agataataca
     2341 ataaccaaca gaaatacgag aatctaaact aatttcgatg attcattttt ctttttaccg
     2401 cgctgcctct tttggcaatt ctttcaccta tattctacct tctctttcct tttgttctaa
     2461 acttattacc agctatctat gtcgaatcaa gaagaaggac ttaaactgtg gggtggcagg
     2521 tttactgggg ctactgaccc cttgatggat ttgtataacg cttccttacc ttacgacaag
     2581 aaaatgtaca aggtggattt agaaggaaca aaagtttaca ctgagggcct ggagaaaatt
     2641 aatttgctaa ctaaagacga actaagtgag attcatcgtg gtctcaaatt gattgaagca
     2701 gagtgggcag aagggaagtt tgttgagaag ccaggggatg aggatattca cactgctaat
     2761 gaacgtcgct tgggtgagtt gattggtcgt ggaatctctg gtaaggttca taccggaagg
     2821 tctagaaatg atcaagttgc cactgatatg cggttgtatg tcagagacaa tctaactcag
     2881 ttggctgact atctgaagca gttcattcaa gtaatcatca agagagctga acaggaaata
     2941 gacgtcttga tgcccggtta tactcacttg caaagagctc aaccaatcag atggtctcac
     3001 tggttgagca tgtatgctac ctatttcact gaagattatg agagactgaa tcaaatcgtt
     3061 aaaaggttga acaaatcccc attgggagct ggagctttgg ctggtcatcc ttatggaatt
     3121 gatcgtgaat acattgctga gagattaggg tttgattctg ttattggtaa ttctttggcc
     3181 gctgtttcag acagagattt tgtagtcgaa accatgttct ggtcttcgtt gtttatgaat
     3241 catatttctc gattctcaga agatttgatc atttactcca ctggagagtt tggatttatc
     3301 aagttggcag atgcttattc tactggatct tctctgatgc ctcaaaaaaa aaacccagac
     3361 tctttggagt tattgagggg taaatctggt agatgttttg gggccttggc tggtttcctc
     3421 atgtctatta agtccattcc gtcaacctat aacaaagata tgcaagagga taaggagcct
     3481 ttatttgata ctctaatcac tgtagagcac tcgattttga tagcatccgg tgtagtttct
     3541 accttgaaca ttgatgccga acgaatgaag aatgctctaa ctatggatat gctggctaca
     3601 gatcttgccg actatttagt tagaagggga gttccattca gagaaactca ccacatttct
     3661 ggtgaatgtg tcagacaagc cgaggagttg aacctttctg gtattgatca gttgtccctc
     3721 gaacaattga aatccattga ctcccgtttt gaggctgatg tggcttcaac gtttgacttt
     3781 gaagccagtg ttgaaaaaag aactgccacc ggaggaactt ctaagactgc tgttttaaag
     3841 caattggatg cactgaatga aaagctagag tcttgactgc aggtcgacag cgtgtttttt
     3901 attccttaag ttacgcaagg tcgaacataa gtacctgtag gatcgtacag gtttacgcaa
     3961 ggtcgaacat aagtacctgt aggatcgtac aggtcgctat tacgccagcg gatccacctg
     4021 taggatcgta caggtgaacc tgtaggatcg tacaggtcaa cctgtaggat cgtacaggta
     4081 cgtatgtgat ccggatgata atgttaagtt acgcaaggtc gaacataagt acctgtagga
     4141 tcgtacaggt ttacgcaagg tcgaacataa gtacctgtag gatcgtacag gtcgctatta
     4201 cgccagcgga tccaactgta aaatcggaca ggtgaaactg taaaatcgga caggtcaaac
     4261 tgtaaaatcg gacaggtacg tatgtgaagc aatatataaa cagaaggaag ctgccctgtc
     4321 ttaaaccttt ttttttatca tcattattag cttactttca taattgcgac tggttccaat
     4381 tgacaagctt ttgattttaa cgacttttaa cgacaacttg agaagatcaa aaaacaacta
     4441 attattcgaa acgactagta tggtttctaa gggtgaagag ttgttcactg gtgttgttcc
     4501 aatcttggtt gagttggacg gtgacgttaa cggacacaag ttctctgttt ctggtgaagg
     4561 tgagggtgac gctacttacg gaaagttgac tttgaagttc atctgtacta ctggtaagtt
     4621 gccagttcca tggccaactt tggttactac tttgacttac ggtgttcagt gtttctccag
     4681 atacccagac cacatgaagc agcacgattt cttcaagtct gctatgccag agggttacgt
     4741 tcaagagaga actatcttct tcaaggacga cggtaactac aagactagag ctgaggttaa
     4801 gttcgagggt gacacattgg ttaacagaat cgagttgaag ggtatcgact tcaaagagga
     4861 cggaaacatc ttgggtcaca agttggagta caactacaac tcccacaacg tttacatcat
     4921 ggctgacaag cagaagaacg gtatcaaggt taacttcaag atcagacaca acatcgagga
     4981 cggttccgtt caattggctg accactacca acagaacact ccaattggtg acggtccagt
     5041 tttgttgcca gacaaccact acttgtccac tcaatccgct ttgtccaagg acccaaacga
     5101 gaagagagat cacatggttt tgttggagtt cgttactgct gctggtatca ctttgggtat
     5161 ggacgagttg tacaagtaac tcgagcctta gacatgactg ttcctcagtt caagttgggc
     5221 acttacgaga agaccggtct tgctagattc taatcaagag gatgtcagaa tgccatttgc
     5281 ctgagagatg caggcttcat ttttgatact tttttatttg taacctatat agtataggat
     5341 tttttttgtc attttgtttc ttctcgtacg agcttgctcc tgatcagcct atctcgcagc
     5401 tgatgaatat cttgtggtag gggtttggga aaatcattcg agtttgatgt ttttcttggt
     5461 atttcccact cctcttcaga gtacagaaga ttaagtgaga cgttcgtttg tgcaagctta
     5521 gatccccggg taccgagctc gaattcgccg taccacttca aaacacccaa gcacagcata
     5581 ctaaatttcc cctctttctt cctctagggt gtcgttaatt acccgtacta aaggtttgga
     5641 aaagaaaaaa gagaccgcct cgtttctttt tcttcgtcga aaaaggcaat aaaaattttt
     5701 atcacgtttc tttttcttga aaattttttt ttttgatttt tttctctttc gatgacctcc
     5761 cattgatatt taagttaata aacggtcttc aatttctcaa gtttcagttt catttttctt
     5821 gttctattac aacttttttt acttcttgct cattagaaag aaagcatagc aatctaatct
     5881 aagttttaat tacaaaatga agaagccaga gttgactgct acttccgttg agaagttctt
     5941 gatcgagaag ttcgactccg tttccgactt gatgcaattg tctgaaggtg aagagtccag
     6001 agctttctcc tttgacgttg gtggtagagg ttacgttttg agagttaatt cctgtgctga
     6061 cggtttctac aaggacagat acgtttacag acacttcgct tccgctgctt tgccaattcc
     6121 agaggttttg gacattggtg agttctccga gtccttgact tactgtatct ccagaagagc
     6181 tcagggagtt acattgcaag acttgccaga aactgagttg ccagctgttt tgcaaccagt
     6241 tgctgaagct atggacgcta ttgctgctgc tgacttgtct caaacttctg gttttggtcc
     6301 attcggtcca cagggtattg gtcagtacac tacttggaga gacttcatct gtgctatcgc
     6361 tgacccacat gtttaccact ggcaaactgt tatggacgat actgtttccg cttccgttgc
     6421 tcaggctttg gacgaattga tgttgtgggc tgaagattgt cctgaggtta gacatttggt
     6481 tcacgctgac ttcggttcca acaacgtttt gactgacaac ggtagaatca ctgctgttat
     6541 cgactggtcc gaagctatgt tcggtgactc tcaatacgag gttgctaaca tcttcttttg
     6601 gagaccatgg ttggcttgta tggaacagca gactagatac ttcgagagaa gacacccaga
     6661 attggctggt tccccaagat tgagagctta catgttgaga atcggtttgg accagttgta
     6721 ccagtccttg gttgatggta acttcgatga tgctgcttgg gctcaaggta gatgtgacgc
     6781 tatcgttaga tccggtgctg gtactgttgg tagaactcag attgctagaa gatccgctgc
     6841 tgtttggact gacggttgtg ttgaagtttt ggctgactcc ggtaacagaa gaccatccac
     6901 tagaccaaga gctaaagagg ttggtagagt ttaagtatta agaacgttat ttatatttca
     6961 aatttttctt ttttttctct acagacgctg tacgcatgta acattatact gaaaaccttg
     7021 cttgagaagg ttttgggacg ctcgaagatc cagctgcatt aatgaatcgg ccaacgcgcg
     7081 gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cggaattcac tggccgtcgt
     7141 tttacaacgt cgtgactggg aaaaccctgg cgttacccaa cttaatcgcc ttgcagcaca
     7201 tccccctttc gccagctggc gtaatagcga agaggcccgc accgatcgcc cttcccaaca
     7261 gttgcgcagc ctgaatggcg aatggcgcct gatgcggtat tttctcctta cgcatctgtg
     7321 cggtatttca caccgcatat ggtgcactct cagtacaatc tgctctgatg ccgcatagtt
     7381 aagccagccc cgacacccgc caacacccgc tgacgcgccc tgacgggctt gtctgctccc
     7441 ggcatccgct tacagacaag ctgtgaccgt ctccgggagc tgcatgtgtc agaggttttc
     7501 accgtcatca ccgaaacgcg cga
//
