LOCUS       Exported                6998 bp DNA     circular SYN 08-APR-2024
DEFINITION  .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6998)
  AUTHORS   Lei Wu
  TITLE     Direct Submission
  JOURNAL   Exported Jun 13, 2024 from SnapGene 6.0.2
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6998
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     2..20
                     /label=DuetUP1
                     /note="color: black; sequence: GGATCTCGACGCTCTCCCT"
     promoter        45..63
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
                     /note="color: #ffffff; direction: RIGHT"
     protein_bind    64..88
                     /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"
     RBS             119..124
                     /note="color: #c0c0c0"
     CDS             132..134
                     /codon_start=1
                     /label=ATG
                     /note="color: #993366"
                     /translation="M"
     CDS             144..161
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /note="color: #cc99b2"
                     /translation="HHHHHH"
     primer_bind     251..270
                     /label=DuetUP2
                     /note="color: black; sequence: TTGTACACGGCCGCATAATC"
     primer_bind     complement(251..270)
                     /label=DuetDOWN
                     /note="color: black; sequence: gattatgcggccgtgtacaa"
     promoter        276..294
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
                     /note="color: #ffffff; direction: RIGHT"
     protein_bind    295..319
                     /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"
     RBS             348..353
                     /note="color: #c0c0c0"
     CDS             362..364
                     /codon_start=1
                     /label=ATG
                     /note="color: #993366"
                     /translation="M"
     primer_bind     370..409
                     /label=UBA1 F
                     /note="color: black; sequence: 
                     tctcaattggatatcgATGCTTCACAAGCGAGCTAGTGAA; added: 
                     2024-04-08"
     CDS             386..3625
                     /codon_start=1
                     /label=AtUBA1
                     /note="color: #00ffff"
                     /translation="MLHKRASEANDKNDNTIIGSDLASSKKRRIDFTESSSDKSSSILA
                     SGSSRGFHGDSVVQQIDMAFGNSNRQEIDEDLHSRQLAVYGRETMRRLFASNVLISGMH
                     GLGAEIAKNLILAGVKSVTLHDERVVELWDLSSNFVFSEDDVGKNRADASVQKLQDLNN
                     AVVVSSLTKSLNKEDLSGFQVVVFSDISMERAIEFDDYCHSHQPPIAFVKADVRGLFGS
                     VFCDFGPEFAVLDVDGEEPHTGIIASISNENQAFISCVDDERLEFEDGDLVVFSEVEGM
                     TELNDGKPRKIKSTRPYSFTLDEDTTNYGTYVKGGIVTQVKQPKLLNFKPLREALKDPG
                     DFLFSDFSKFDRPPLLHLAFQALDHFKAEAGRFPVAGSEEDAQKLISIATAINTGQGDL
                     KVENVDQKLLRHFSFGAKAVLNPMAAMFGGIVGQEVVKACSGKFHPLFQFFYFDSVESL
                     PSEPVDSSDFAPRNSRYDAQISVFGAKFQKKLEDAKVFTVGSGALGCEFLKNLALMGVS
                     CGSQGKLTVTDDDIIEKSNLSRQFLFRDWNIGQAKSTVAASAAAVINPRFNIEALQNRV
                     GAETENVFDDAFWENLTVVVNALDNVNARLYVDSRCLYFQKPLLESGTLGTKCNTQSVI
                     PHLTENYGASRDPPEKQAPMCTVHSFPHNIDHCLTWARSEFEGLLEKTPAEVNAYLSSP
                     VEYTNSMMSAGDAQARDTLERIVECLEKEKCETFQDCLTWARLRFEDYFVNRVKQLIYT
                     FPEDAATSTGAPFWSAPKRFPRPLQYSSSDPSLLNFITATAILRAETFGIPIPEWTKNP
                     KEAAEAVDRVIVPDFEPRQDAKIVTDEKATTLTTASVDDAAVIDDLIAKIDQCRHNLSP
                     DFRMKPIQFEKDDDTNYHMDVIAGLANMRARNYSIPEVDKLKAKFIAGRIIPAIATSTA
                     MATGLVCLELYKVLDGGHKVEAYRNTFANLALPLFSMAEPLPPKVVKHRDMAWTVWDRW
                     VLKGNPTLREVLQWLEDKGLSAYSISCGSCLLFNSMFTRHKERMDKKVVDLARDVAKVE
                     LPPYRNHLDVVVACEDEDDNDVDIPLVSIYFR"
     primer_bind     complement(3601..3640)
                     /label=UBA1 R
                     /note="color: black; sequence: 
                     agactcgagggtaccCCTGAAGTAGATAGAGACGAGAGGA; added: 
                     2024-04-08"
     CDS             3644..3688
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S
                     /note="color: #cc99b2"
                     /translation="KETAAAKFERQHMDS"
     terminator      3740..3787
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
                     /note="color: #ffffff"
     promoter        3854..3872
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
                     /note="color: #ffffff; direction: RIGHT"
     protein_bind    3873..3897
                     /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"
     CDS             complement(3958..4749)
                     /codon_start=1
                     /gene="aadA"
                     /product="aminoglycoside adenylyltransferase (Murphy, 
                     1985)"
                     /label=SmR
                     /note="confers resistance to spectinomycin and 
                     streptomycin"
                     /note="color: #ccffcc"
                     /translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
                     SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
                     RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
                     EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
                     VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK"
     promoter        complement(4750..4841)
                     /gene="bla"
                     /label=AmpR promoter
                     /note="color: #ffffff; direction: LEFT"
     rep_origin      complement(4889..5627)
                     /direction=LEFT
                     /label=CloDF13 ori
                     /note="Plasmids containing the CloDF13 (CDF) origin of 
                     replication can be propagated in E. coli cells that contain
                     additional plasmids with compatible origins."
                     /note="color: #ffff00"
     CDS             complement(5837..6919)
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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: #993366"
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     promoter        complement(6920..6997)
                     /gene="lacI"
                     /label=lacI promoter
                     /note="color: #ffffff; direction: LEFT"
ORIGIN
        1 gggatctcga cgctctccct tatgcgactc ctgcattagg aaattaatac gactcactat
       61 aggggaattg tgagcggata acaattcccc tgtagaaata attttgttta actttaataa
      121 ggagatatac catgggcagc agccatcacc atcatcacca cagccaggga tccgaattcg
      181 agctcggcgc gcctgcaggt cgacaagctt gcggccgcat aatgcttaag tcgaacagaa
      241 agtaatcgta ttgtacacgg ccgcataatc gaaattaata cgactcacta taggggaatt
      301 gtgagcggat aacaattccc catcttagta tattagttaa gtataagaag gagatataca
      361 tatggcagat ctcaattgga tatcgatgct tcacaagcga gctagtgaag ctaacgataa
      421 gaacgataat accatcatcg gttccgattt agcatcatct aagaaacgcc gtatcgattt
      481 cactgaatct tcgtctgata agtcttcttc gattttagct tctggtagta gcaggggttt
      541 tcacggcgat agcgtcgttc agcaaatcga catggctttt ggtaattcga accgtcagga
      601 gattgatgaa gatctgcaca gtcgccagct cgccgtctat ggtcgtgaga ctatgaggcg
      661 tctctttgct tccaatgttc tcatctcggg gatgcacggt cttggtgccg agattgccaa
      721 gaatcttata cttgctggtg tgaagtctgt gaccttgcat gatgaaagag ttgtagagct
      781 atgggactta tcaagcaact ttgttttctc ggaggatgat gttggcaaga atagagcaga
      841 tgcttctgtt cagaagttac aggatcttaa caatgctgtg gttgtctcta gcttgaccaa
      901 aagcttaaac aaagaggatc tttctggctt ccaggttgtt gtattctctg acataagcat
      961 ggaaagagca atcgaatttg atgactattg tcacagccat cagcctccta tcgcttttgt
     1021 caaggctgat gtcaggggtc ttttcggctc tgtcttttgt gattttgggc ctgaatttgc
     1081 ggttcttgat gttgatggag aggaacccca tacaggcatc attgcgtcta tctctaatga
     1141 gaaccaggca tttatttcct gtgtcgacga tgagaggctt gagtttgaag atggtgacct
     1201 agttgttttc tcagaagttg agggtatgac agaattgaat gatgggaaac caaggaagat
     1261 aaaaagtaca cggccatatt cattcaccct tgatgaggac actactaact atggaacata
     1321 tgtgaagggt ggtattgtca ctcaagtgaa acagccaaag ttactgaatt tcaagccatt
     1381 gagggaagcg cttaaggatc caggggattt tctatttagt gatttctcta agtttgatcg
     1441 gcctccactc cttcacttag cattccaggc acttgatcat tttaaagctg aagctggcag
     1501 atttcctgtt gctggctcag aagaggacgc tcagaagctt atatcgattg ccacagccat
     1561 caataccggg cagggtgatt taaaggtgga gaatgttgat cagaagcttc taagacactt
     1621 ttcctttgga gccaaggctg tcctgaatcc tatggctgca atgtttggcg gtattgttgg
     1681 acaggaggtt gttaaagctt gctctggaaa atttcatcct ctcttccagt ttttctactt
     1741 tgattcagtg gagtcactcc cctctgaacc tgtggactcg agtgactttg caccaagaaa
     1801 cagccggtat gatgcccaaa tatctgtatt tggggccaag ttccagaaga aacttgaaga
     1861 tgctaaagtt ttcacagtag ggtctggtgc tcttggctgt gagttcttga aaaatttggc
     1921 tctgatgggg gtttcttgtg gaagccaagg gaagctgaca gtgacagatg atgatatcat
     1981 tgagaagagt aacctcagtc gtcaatttct gttccgtgac tggaacattg gacaggctaa
     2041 atccacagtt gctgcttcgg ctgctgctgt tataaatcct cgcttcaaca ttgaggccct
     2101 gcaaaaccgt gttggcgcgg agactgagaa tgtatttgac gatgccttct gggagaactt
     2161 aactgttgtc gtcaatgcac tggataatgt taatgcgagg ctctatgttg attcgaggtg
     2221 cttgtatttc cagaagcctc tccttgagtc tggaactctt ggtacaaagt gcaacacaca
     2281 gtcggttatc ccacatctaa ctgaaaatta cggtgcctca agggacccac cagagaaaca
     2341 ggcccccatg tgtacagtgc attccttccc gcataacatt gatcactgtt taacttgggc
     2401 tcgctctgag tttgagggtc tgctggagaa gactcccgct gaagtgaatg catatctttc
     2461 tagcccagtt gagtacacta actcaatgat gagtgctggt gatgctcagg cgagggacac
     2521 attggagagg attgttgagt gccttgaaaa ggagaagtgt gagaccttcc aagactgctt
     2581 aacctgggca cgactgaggt ttgaggatta ctttgtgaac cgtgtcaagc aattgatata
     2641 cacatttcct gaagatgctg cgactagtac cggagctcca ttctggtctg ctccaaaaag
     2701 attccctcgt ccactccagt actcgtcgtc cgacccaagt ctccttaact ttatcacagc
     2761 gactgctatt ttacgagcag agacatttgg gatcccgata cctgagtgga ctaagaaccc
     2821 aaaggaagca gcagaagctg ttgacagagt gatagtccca gactttgagc caaggcaaga
     2881 tgcaaagatt gtgactgatg agaaagctac cactttaacc actgcttcgg tggacgacgc
     2941 tgcagtcatc gacgatctca ttgctaagat cgaccagtgt aggcataact tgtcgccaga
     3001 cttcaggatg aaacctattc agttcgaaaa ggatgatgat acaaactatc acatggatgt
     3061 gatagcagga ctagccaaca tgagggcgag aaattacagc atccctgaag tggacaagct
     3121 gaaagcaaag ttcatagcag ggagaatcat cccagccatt gcgacttcaa cagccatggc
     3181 tactgggctg gtctgcctcg agctgtacaa ggtccttgat ggaggacata aagtggaagc
     3241 ttacagaaac acatttgcca acctggcact tccactattc tccatggctg aaccacttcc
     3301 accaaaggtg gtgaagcacc gtgacatggc ttggacagtt tgggacagat gggttctcaa
     3361 gggaaaccca actcttcgtg aggtgttgca gtggctggag gacaaagggc ttagcgcata
     3421 cagcatctct tgtggaagct gtcttctgtt caacagtatg ttcacaaggc acaaggagag
     3481 gatggacaag aaggttgtgg atctcgctcg ggatgttgcg aaagtcgagt taccgcctta
     3541 tcgcaaccat cttgatgtag tggtggcttg tgaggatgaa gatgacaatg acgtcgatat
     3601 tcctctcgtc tctatctact tcaggggtac cctcgagtct ggtaaagaaa ccgctgctgc
     3661 gaaatttgaa cgccagcaca tggactcgtc tactagcgca gcttaattaa cctaggctgc
     3721 tgccaccgct gagcaataac tagcataacc ccttggggcc tctaaacggg tcttgagggg
     3781 ttttttgctg aaacctcagg catttgagaa gcacacggtc acactgcttc cggtagtcaa
     3841 taaaccggta aaccagcaat agacataagc ggctatttaa cgaccctgcc ctgaaccgac
     3901 gaccgggtca tcgtggccgg atcttgcggc ccctcggctt gaacgaattg ttagacatta
     3961 tttgccgact accttggtga tctcgccttt cacgtagtgg acaaattctt ccaactgatc
     4021 tgcgcgcgag gccaagcgat cttcttcttg tccaagataa gcctgtctag cttcaagtat
     4081 gacgggctga tactgggccg gcaggcgctc cattgcccag tcggcagcga catccttcgg
     4141 cgcgattttg ccggttactg cgctgtacca aatgcgggac aacgtaagca ctacatttcg
     4201 ctcatcgcca gcccagtcgg gcggcgagtt ccatagcgtt aaggtttcat ttagcgcctc
     4261 aaatagatcc tgttcaggaa ccggatcaaa gagttcctcc gccgctggac ctaccaaggc
     4321 aacgctatgt tctcttgctt ttgtcagcaa gatagccaga tcaatgtcga tcgtggctgg
     4381 ctcgaagata cctgcaagaa tgtcattgcg ctgccattct ccaaattgca gttcgcgctt
     4441 agctggataa cgccacggaa tgatgtcgtc gtgcacaaca atggtgactt ctacagcgcg
     4501 gagaatctcg ctctctccag gggaagccga agtttccaaa aggtcgttga tcaaagctcg
     4561 ccgcgttgtt tcatcaagcc ttacggtcac cgtaaccagc aaatcaatat cactgtgtgg
     4621 cttcaggccg ccatccactg cggagccgta caaatgtacg gccagcaacg tcggttcgag
     4681 atggcgctcg atgacgccaa ctacctctga tagttgagtc gatacttcgg cgatcaccgc
     4741 ttccctcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat
     4801 gagcggatac atatttgaat gtatttagaa aaataaacaa atagctagct cactcggtcg
     4861 ctacgctccg ggcgtgagac tgcggcgggc gctgcggaca catacaaagt tacccacaga
     4921 ttccgtggat aagcagggga ctaacatgtg aggcaaaaca gcagggccgc gccggtggcg
     4981 tttttccata ggctccgccc tcctgccaga gttcacataa acagacgctt ttccggtgca
     5041 tctgtgggag ccgtgaggct caaccatgaa tctgacagta cgggcgaaac ccgacaggac
     5101 ttaaagatcc ccaccgtttc cggcgggtcg ctccctcttg cgctctcctg ttccgaccct
     5161 gccgtttacc ggatacctgt tccgcctttc tcccttacgg gaagtgtggc gctttctcat
     5221 agctcacaca ctggtatctc ggctcggtgt aggtcgttcg ctccaagctg ggctgtaagc
     5281 aagaactccc cgttcagccc gactgctgcg ccttatccgg taactgttca cttgagtcca
     5341 acccggaaaa gcacggtaaa acgccactgg cagcagccat tggtaactgg gagttcgcag
     5401 aggatttgtt tagctaaaca cgcggttgct cttgaagtgt gcgccaaagt ccggctacac
     5461 tggaaggaca gatttggttg ctgtgctctg cgaaagccag ttaccacggt taagcagttc
     5521 cccaactgac ttaaccttcg atcaaaccac ctccccaggt ggttttttcg tttacagggc
     5581 aaaagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctactgaac
     5641 cgctctagat ttcagtgcaa tttatctctt caaatgtagc acctgaagtc agccccatac
     5701 gatataagtt gtaattctca tgttagtcat gccccgcgcc caccggaagg agctgactgg
     5761 gttgaaggct ctcaagggca tcggtcgaga tcccggtgcc taatgagtga gctaacttac
     5821 attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca
     5881 ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgcc agggtggttt
     5941 ttcttttcac cagtgagacg ggcaacagct gattgccctt caccgcctgg ccctgagaga
     6001 gttgcagcaa gcggtccacg ctggtttgcc ccagcaggcg aaaatcctgt ttgatggtgg
     6061 ttaacggcgg gatataacat gagctgtctt cggtatcgtc gtatcccact accgagatgt
     6121 ccgcaccaac gcgcagcccg gactcggtaa tggcgcgcat tgcgcccagc gccatctgat
     6181 cgttggcaac cagcatcgca gtgggaacga tgccctcatt cagcatttgc atggtttgtt
     6241 gaaaaccgga catggcactc cagtcgcctt cccgttccgc tatcggctga atttgattgc
     6301 gagtgagata tttatgccag ccagccagac gcagacgcgc cgagacagaa cttaatgggc
     6361 ccgctaacag cgcgatttgc tggtgaccca atgcgaccag atgctccacg cccagtcgcg
     6421 taccgtcttc atgggagaaa ataatactgt tgatgggtgt ctggtcagag acatcaagaa
     6481 ataacgccgg aacattagtg caggcagctt ccacagcaat ggcatcctgg tcatccagcg
     6541 gatagttaat gatcagccca ctgacgcgtt gcgcgagaag attgtgcacc gccgctttac
     6601 aggcttcgac gccgcttcgt tctaccatcg acaccaccac gctggcaccc agttgatcgg
     6661 cgcgagattt aatcgccgcg acaatttgcg acggcgcgtg cagggccaga ctggaggtgg
     6721 caacgccaat cagcaacgac tgtttgcccg ccagttgttg tgccacgcgg ttgggaatgt
     6781 aattcagctc cgccatcgcc gcttccactt tttcccgcgt tttcgcagaa acgtggctgg
     6841 cctggttcac cacgcgggaa acggtctgat aagagacacc ggcatactct gcgacatcgt
     6901 ataacgttac tggtttcaca ttcaccaccc tgaattgact ctcttccggg cgctatcatg
     6961 ccataccgcg aaaggttttg cgccattcga tggtgtcc
//
