LOCUS       Exported                5993 bp ds-DNA     circular SYN 20-SEP-2021
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    pGEX4T3-GST-Polß(K206A:K244A:T304I)
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5993)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Monday, Sep 27, 2021 from SnapGene 3.3.3
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..5993
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     promoter        183..211
                     /note="tac promoter"
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     misc_feature    184..211
                     /label=tac promoter
                     /note="tac promoter"
     protein_bind    219..235
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="lac operator"
                     /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)."
     misc_feature    258..935
                     /label=GST
                     /note="GST"
     CDS             258..911
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /note="GST"
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     misc_feature    918..935
                     /label=Thrombin
                     /note="Thrombin"
     CDS             918..935
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /note="thrombin site"
                     /translation="LVPRGS"
     misc_feature    936..956
                     /label=Tev site
                     /note="Tev site"
     CDS             936..956
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /note="TEV site"
                     /translation="ENLYFQG"
     misc_feature    957
                     /label=New Feature(1)
                     /note="New Feature"
     misc_feature    958..974
                     /label=GST(1)
                     /note="GST"
     misc_feature    959..964
                     /label=EcoRI
                     /note="EcoRI"
     misc_feature    965..969
                     /label=SmaI
                     /note="SmaI"
     misc_feature    join(975..1589,1592..1703,1706..1884,1886..1982)
                     /label=PolB
                     /note="PolB"
     misc_feature    join(975..1589,1592..1703,1706..1884,1886..1982)
                     /label=Polb(WT)
                     /note="Polb(WT)"
     misc_feature    1590..1592
                     /label=K206A
                     /note="K206A"
     misc_feature    1704..1706
                     /label=K244A
                     /note="K244A"
     misc_feature    1884..1886
                     /label=T304I
                     /note="T304I"
     misc_feature    1983..2000
                     /label=GST(2)
                     /note="GST"
     misc_feature    2043..2064
                     /label=pGEX 3 primer
                     /note="pGEX 3 primer"
     promoter        2296..2400
                     /gene="bla"
                     /note="AmpR promoter"
     misc_feature    2331..2358
                     /label=AmpR promoter
                     /note="AmpR promoter"
     misc_feature    2401..3261
                     /label=Ampicillin
                     /note="Ampicillin"
     CDS             2401..3261
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /note="AmpR"
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      3432..4020
                     /direction=RIGHT
                     /note="ori"
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     promoter        4264..4341
                     /gene="lacI (mutant)"
                     /note="lacIq promoter"
                     /note="In the lacIq allele, a single base change in the 
                     promoter boosts expression of the lacI gene about 10-fold."
     misc_feature    4333
                     /label=New Feature
                     /note="New Feature"
     misc_feature    4334..5424
                     /label=lacI
                     /note="lacI"
     CDS             4342..5424
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /note="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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     protein_bind    5437..5458
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding site"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        5473..5503
                     /note="lac promoter"
                     /note="promoter for the E. coli lac operon"
     protein_bind    5511..5527
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="lac operator"
                     /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     5535..5551
                     /note="M13 rev"
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             5547..81
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /note="lacZ-alpha"
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGITLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(5567..5583)
                     /note="M13 fwd"
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 acgttatcga ctgcacggtg caccaatgct tctggcgtca ggcagccatc ggaagctgtg
       61 gtatggctgt gcaggtcgta aatcactgca taattcgtgt cgctcaaggc gcactcccgt
      121 tctggataat gttttttgcg ccgacatcat aacggttctg gcaaatattc tgaaatgagc
      181 tgttgacaat taatcatcgg ctcgtataat gtgtggaatt gtgagcggat aacaatttca
      241 cacaggaaac agtattcatg tcccctatac taggttattg gaaaattaag ggccttgtgc
      301 aacccactcg acttcttttg gaatatcttg aagaaaaata tgaagagcat ttgtatgagc
      361 gcgatgaagg tgataaatgg cgaaacaaaa agtttgaatt gggtttggag tttcccaatc
      421 ttccttatta tattgatggt gatgttaaat taacacagtc tatggccatc atacgttata
      481 tagctgacaa gcacaacatg ttgggtggtt gtccaaaaga gcgtgcagag atttcaatgc
      541 ttgaaggagc ggttttggat attagatacg gtgtttcgag aattgcatat agtaaagact
      601 ttgaaactct caaagttgat tttcttagca agctacctga aatgctgaaa atgttcgaag
      661 atcgtttatg tcataaaaca tatttaaatg gtgatcatgt aacccatcct gacttcatgt
      721 tgtatgacgc tcttgatgtt gttttataca tggacccaat gtgcctggat gcgttcccaa
      781 aattagtttg ttttaaaaaa cgtattgaag ctatcccaca aattgataag tacttgaaat
      841 ccagcaagta tatagcatgg cctttgcagg gctggcaagc cacgtttggt ggtggcgacc
      901 atcctccaaa atcggatctg gttccgcgtg gatccgaaaa cctgtatttt cagggctcga
      961 attcccgggt cgacatgagc aaacggaagg cgccgcagga gactctcaac gggggaatca
     1021 ccgacatgct cacagaactc gcaaactttg agaagaacgt gagccaagct atccacaagt
     1081 acaatgctta cagaaaagca gcatctgtta tagcaaaata cccacacaaa ataaagagtg
     1141 gagctgaagc taagaaattg cctggagtag gaacaaaaat tgctgaaaag attgatgagt
     1201 ttttagcaac tggaaaatta cgtaaactgg aaaagattcg gcaggatgat acgagttcat
     1261 ccatcaattt cctgactcga gttagtggca ttggtccatc tgctgcaagg aagtttgtag
     1321 atgaaggaat taaaacacta gaagatctca gaaaaaatga agataaattg aaccatcatc
     1381 agcgaattgg gctgaaatat tttggggact ttgaaaaaag aattcctcgt gaagagatgt
     1441 tacaaatgca agatattgta ctaaatgaag ttaaaaaagt ggattctgaa tacattgcta
     1501 cagtctgtgg cagtttcaga agaggtgcag agtccagtgg tgacatggat gttctcctga
     1561 cccatcccag cttcacttca gaatcaaccg cacagccaaa actgttacat caggttgtgg
     1621 agcagttaca aaaggttcat tttatcacag ataccctgtc aaagggtgag acaaagttca
     1681 tgggtgtttg ccagcttccc agtgcaaatg atgaaaaaga atatccacac agaagaattg
     1741 atatcaggtt gatacccaaa gatcagtatt actgtggtgt tctctatttc actgggagtg
     1801 atattttcaa taagaatatg agggctcatg ccctagaaaa gggtttcaca atcaatgagt
     1861 acaccatccg tcccttggga gtcattggag ttgcaggaga acccctgcca gtggatagtg
     1921 aaaaagacat ctttgattac atccagtgga aataccggga acccaaggac cggagcgaat
     1981 gagcggccgc atcgtgactg actgacgatc tgcctcgcgc gtttcggtga tgacggtgaa
     2041 aacctctgac acatgcagct cccggagacg gtcacagctt gtctgtaagc ggatgccggg
     2101 agcagacaag cccgtcaggg cgcgtcagcg ggtgttggcg ggtgtcgggg cgcagccatg
     2161 acccagtcac gtagcgatag cggagtgtat aattcttgaa gacgaaaggg cctcgtgata
     2221 cgcctatttt tataggttaa tgtcatgata ataatggttt cttagacgtc aggtggcact
     2281 tttcggggaa atgtgcgcgg aacccctatt tgtttatttt tctaaataca ttcaaatatg
     2341 tatccgctca tgagacaata accctgataa atgcttcaat aatattgaaa aaggaagagt
     2401 atgagtattc aacatttccg tgtcgccctt attccctttt ttgcggcatt ttgccttcct
     2461 gtttttgctc acccagaaac gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca
     2521 cgagtgggtt acatcgaact ggatctcaac agcggtaaga tccttgagag ttttcgcccc
     2581 gaagaacgtt ttccaatgat gagcactttt aaagttctgc tatgtggcgc ggtattatcc
     2641 cgtgttgacg ccgggcaaga gcaactcggt cgccgcatac actattctca gaatgacttg
     2701 gttgagtact caccagtcac agaaaagcat cttacggatg gcatgacagt aagagaatta
     2761 tgcagtgctg ccataaccat gagtgataac actgcggcca acttacttct gacaacgatc
     2821 ggaggaccga aggagctaac cgcttttttg cacaacatgg gggatcatgt aactcgcctt
     2881 gatcgttggg aaccggagct gaatgaagcc ataccaaacg acgagcgtga caccacgatg
     2941 cctgcagcaa tggcaacaac gttgcgcaaa ctattaactg gcgaactact tactctagct
     3001 tcccggcaac aattaataga ctggatggag gcggataaag ttgcaggacc acttctgcgc
     3061 tcggcccttc cggctggctg gtttattgct gataaatctg gagccggtga gcgtgggtct
     3121 cgcggtatca ttgcagcact ggggccagat ggtaagccct cccgtatcgt agttatctac
     3181 acgacgggga gtcaggcaac tatggatgaa cgaaatagac agatcgctga gataggtgcc
     3241 tcactgatta agcattggta actgtcagac caagtttact catatatact ttagattgat
     3301 ttaaaacttc atttttaatt taaaaggatc taggtgaaga tcctttttga taatctcatg
     3361 accaaaatcc cttaacgtga gttttcgttc cactgagcgt cagaccccgt agaaaagatc
     3421 aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa
     3481 ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct ttttccgaag
     3541 gtaactggct tcagcagagc gcagatacca aatactgtcc ttctagtgta gccgtagtta
     3601 ggccaccact tcaagaactc tgtagcaccg cctacatacc tcgctctgct aatcctgtta
     3661 ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg ggttggactc aagacgatag
     3721 ttaccggata aggcgcagcg gtcgggctga acggggggtt cgtgcacaca gcccagcttg
     3781 gagcgaacga cctacaccga actgagatac ctacagcgtg agctatgaga aagcgccacg
     3841 cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg aacaggagag
     3901 cgcacgaggg agcttccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc
     3961 cacctctgac ttgagcgtcg atttttgtga tgctcgtcag gggggcggag cctatggaaa
     4021 aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg
     4081 ttctttcctg cgttatcccc tgattctgtg gataaccgta ttaccgcctt tgagtgagct
     4141 gataccgctc gccgcagccg aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa
     4201 gagcgcctga tgcggtattt tctccttacg catctgtgcg gtatttcaca ccgcataaat
     4261 tccgacacca tcgaatggtg caaaaccttt cgcggtatgg catgatagcg cccggaagag
     4321 agtcaattca gggtggtgaa tgtgaaacca gtaacgttat acgatgtcgc agagtatgcc
     4381 ggtgtctctt atcagaccgt ttcccgcgtg gtgaaccagg ccagccacgt ttctgcgaaa
     4441 acgcgggaaa aagtggaagc ggcgatggcg gagctgaatt acattcccaa ccgcgtggca
     4501 caacaactgg cgggcaaaca gtcgttgctg attggcgttg ccacctccag tctggccctg
     4561 cacgcgccgt cgcaaattgt cgcggcgatt aaatctcgcg ccgatcaact gggtgccagc
     4621 gtggtggtgt cgatggtaga acgaagcggc gtcgaagcct gtaaagcggc ggtgcacaat
     4681 cttctcgcgc aacgcgtcag tgggctgatc attaactatc cgctggatga ccaggatgcc
     4741 attgctgtgg aagctgcctg cactaatgtt ccggcgttat ttcttgatgt ctctgaccag
     4801 acacccatca acagtattat tttctcccat gaagacggta cgcgactggg cgtggagcat
     4861 ctggtcgcat tgggtcacca gcaaatcgcg ctgttagcgg gcccattaag ttctgtctcg
     4921 gcgcgtctgc gtctggctgg ctggcataaa tatctcactc gcaatcaaat tcagccgata
     4981 gcggaacggg aaggcgactg gagtgccatg tccggttttc aacaaaccat gcaaatgctg
     5041 aatgagggca tcgttcccac tgcgatgctg gttgccaacg atcagatggc gctgggcgca
     5101 atgcgcgcca ttaccgagtc cgggctgcgc gttggtgcgg atatctcggt agtgggatac
     5161 gacgataccg aagacagctc atgttatatc ccgccgttaa ccaccatcaa acaggatttt
     5221 cgcctgctgg ggcaaaccag cgtggaccgc ttgctgcaac tctctcaggg ccaggcggtg
     5281 aagggcaatc agctgttgcc cgtctcactg gtgaaaagaa aaaccaccct ggcgcccaat
     5341 acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc acgacaggtt
     5401 tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc tcactcatta
     5461 ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa ttgtgagcgg
     5521 ataacaattt cacacaggaa acagctatga ccatgattac ggattcactg gccgtcgttt
     5581 tacaacgtcg tgactgggaa aaccctggcg ttacccaact taatcgcctt gcagcacatc
     5641 cccctttcgc cagctggcgt aatagcgaag aggcccgcac cgatcgccct tcccaacagt
     5701 tgcgcagcct gaatggcgaa tggcgctttg cctggtttcc ggcaccagaa gcggtgccgg
     5761 aaagctggct ggagtgcgat cttcctgagg ccgatactgt cgtcgtcccc tcaaactggc
     5821 agatgcacgg ttacgatgcg cccatctaca ccaacgtaac ctatcccatt acggtcaatc
     5881 cgccgtttgt tcccacggag aatccgacgg gttgttactc gctcacattt aatgttgatg
     5941 aaagctggct acaggaaggc cagacgcgaa ttatttttga tggcgttgga att
//
