LOCUS       Exported                7176 bp ds-DNA    circular SYN 04-JAN-2019
DEFINITION  pET-21 b (+).
ACCESSION   .
VERSION     .
KEYWORDS    18 - MHET6CCPET
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7176)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jan 4, 2019 from SnapGene 2.3.2
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7176
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     source          1..5204
                     /organism="pET-21 b (+)"
                     /mol_type="other DNA"
                     /label="source:pET-21 b (+)"
                     /note="source:pET-21 b"
     source          5184..7049
                     /label="gBlock: CJ_MHETase_opt_Ec"
                     /note="gBlock: CJ_MHETase_opt_Ec"
     source          7014..7176
                     /organism="pET-21 b (+)"
                     /mol_type="other DNA"
                     /label="source:pET-21 b (+)(1)"
                     /note="source:pET-21 b"
     promoter        529..557
                     /label="AmpR_promoter"
                     /note="AmpR_promoter"
     CDS             599..1459
                     /codon_start=1
                     /label="ORF frame 2(1)"
                     /note="ORF frame 2"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     gene            599..1459
                     /gene="Ampicillin"
                     /label="Ampicillin"
                     /note="Ampicillin"
     rep_origin      1614..2233
                     /label="pBR322_origin"
                     /note="pBR322_origin"
     misc_feature    2632..2654
                     /label="pGEX_3_primer"
                     /note="pGEX_3_primer"
     misc_feature    complement(2648..2839)
                     /label="ROP"
                     /note="ROP"
     gene            complement(3314..3589)
                     /gene="tet (576 - 851)"
                     /label="tet (576 - 851)"
                     /note="tet"
     misc_feature    complement(3648..4739)
                     /label="lacI"
                     /note="lacI"
     CDS             complement(3648..4607)
                     /codon_start=1
                     /label="ORF frame 2"
                     /note="ORF frame 2"
                     /translation="MAELNYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSR
                     ADQLGASVVVSMVERSGVEACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPA
                     LFLDVSDQTPINSIIFSHEDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKY
                     LTRNQIQPIAEREGDWSAMSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLR
                     VGADISVVGYDDTEDSSCYIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVS
                     LVKRKTTLAPNTQTASPRALADSLMQLARQVSRLESGQ"
     CDS             complement(4480..4980)
                     /codon_start=1
                     /label="ORF frame 1"
                     /note="ORF frame 1"
                     /translation="MSACFGVGMVAGPVAGGLLGAISLHAPFLAAAVLNGLNLLLGCFL
                     MQESHKGERRDPGHHRMAQNLSRYGMIAPGRESIQGGECETSNVIRCRRVCRCLLSDRF
                     PRGEPGQPRFCENAGKSGSGDGGAELHSQPRGTTTGGQTVVADWRCHLQSGPARAVANC
                     RGD"
     gene            complement(4822..5085)
                     /gene="tet (300 - 563)"
                     /label="tet (300 - 563)"
                     /note="tet"
     misc_feature    5014..5033
                     /label="pBRrevBam_primer"
                     /note="pBRrevBam_primer"
     primer_bind     complement(5117..5136)
                     /label="T7 primer"
                     /note="T7 primer"
     promoter        5117..5135
                     /label="T7_promoter"
                     /note="T7_promoter"
     misc_feature    5135..5162
                     /label="lacO"
                     /note="lacO"
     misc_feature    5180..5196
                     /label="T7_transl_en_RBS"
                     /note="T7_transl_en_RBS"
     misc_feature    5204
                     /label="Xpress_fwd_primer"
                     /note="Xpress_fwd_primer"
     CDS             5205..7013
                     /label="MHETase gene opt E.c."
                     /note="MHETase gene opt E.c"
     primer_bind     5768..5786
                     /label="oCJ668"
                     /note="oCJ668"
     primer_bind     complement(6642..6668)
                     /note="oCJ780"
     primer_bind     6669..6693
                     /note="oCJ779"
     variation       6669..6671
                     /note="Gly489Cys Mutation"
     primer_bind     complement(6770..6794)
                     /note="oCJ782"
     variation       6792..6794
                     /note="Ser530Cys Mutation"
     primer_bind     6795..6812
                     /note="oCJ781"
     misc_signal     7020..7037
                     /label="His Tag"
                     /note="His Tag"
     terminator      7048..7176
                     /label="T7_terminator"
                     /note="T7_terminator"
     primer_bind     complement(7090..7108)
                     /label="T7 Term primer"
                     /note="T7 Term primer"
     misc_feature    complement(7090..7108)
                     /label="T7_Terminal_primer"
                     /note="T7_Terminal_primer"
ORIGIN
        1 tggcgaatgg gacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg
       61 cagcgtgacc gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc
      121 ctttctcgcc acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg
      181 gttccgattt agtgctttac ggcacctcga ccccaaaaaa cttgattagg gtgatggttc
      241 acgtagtggg ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt
      301 ctttaatagt ggactcttgt tccaaactgg aacaacactc aaccctatct cggtctattc
      361 ttttgattta taagggattt tgccgatttc ggcctattgg ttaaaaaatg agctgattta
      421 acaaaaattt aacgcgaatt ttaacaaaat attaacgttt acaatttcag gtggcacttt
      481 tcggggaaat gtgcgcggaa cccctatttg tttatttttc taaatacatt caaatatgta
      541 tccgctcatg agacaataac cctgataaat gcttcaataa tattgaaaaa ggaagagtat
      601 gagtattcaa catttccgtg tcgcccttat tccctttttt gcggcatttt gccttcctgt
      661 ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg
      721 agtgggttac atcgaactgg atctcaacag cggtaagatc cttgagagtt ttcgccccga
      781 agaacgtttt ccaatgatga gcacttttaa agttctgcta tgtggcgcgg tattatcccg
      841 tattgacgcc gggcaagagc aactcggtcg ccgcatacac tattctcaga atgacttggt
      901 tgagtactca ccagtcacag aaaagcatct tacggatggc atgacagtaa gagaattatg
      961 cagtgctgcc ataaccatga gtgataacac tgcggccaac ttacttctga caacgatcgg
     1021 aggaccgaag gagctaaccg cttttttgca caacatgggg gatcatgtaa ctcgccttga
     1081 tcgttgggaa ccggagctga atgaagccat accaaacgac gagcgtgaca ccacgatgcc
     1141 tgcagcaatg gcaacaacgt tgcgcaaact attaactggc gaactactta ctctagcttc
     1201 ccggcaacaa ttaatagact ggatggaggc ggataaagtt gcaggaccac ttctgcgctc
     1261 ggcccttccg gctggctggt ttattgctga taaatctgga gccggtgagc gtgggtctcg
     1321 cggtatcatt gcagcactgg ggccagatgg taagccctcc cgtatcgtag ttatctacac
     1381 gacggggagt caggcaacta tggatgaacg aaatagacag atcgctgaga taggtgcctc
     1441 actgattaag cattggtaac tgtcagacca agtttactca tatatacttt agattgattt
     1501 aaaacttcat ttttaattta aaaggatcta ggtgaagatc ctttttgata atctcatgac
     1561 caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa
     1621 aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc
     1681 accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt
     1741 aactggcttc agcagagcgc agataccaaa tactgtcctt ctagtgtagc cgtagttagg
     1801 ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc
     1861 agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt
     1921 accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga
     1981 gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct
     2041 tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg
     2101 cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca
     2161 cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa
     2221 cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgtt
     2281 ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga
     2341 taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga
     2401 gcgcctgatg cggtattttc tccttacgca tctgtgcggt atttcacacc gcatatatgg
     2461 tgcactctca gtacaatctg ctctgatgcc gcatagttaa gccagtatac actccgctat
     2521 cgctacgtga ctgggtcatg gctgcgcccc gacacccgcc aacacccgct gacgcgccct
     2581 gacgggcttg tctgctcccg gcatccgctt acagacaagc tgtgaccgtc tccgggagct
     2641 gcatgtgtca gaggttttca ccgtcatcac cgaaacgcgc gaggcagctg cggtaaagct
     2701 catcagcgtg gtcgtgaagc gattcacaga tgtctgcctg ttcatccgcg tccagctcgt
     2761 tgagtttctc cagaagcgtt aatgtctggc ttctgataaa gcgggccatg ttaagggcgg
     2821 ttttttcctg tttggtcact gatgcctccg tgtaaggggg atttctgttc atgggggtaa
     2881 tgataccgat gaaacgagag aggatgctca cgatacgggt tactgatgat gaacatgccc
     2941 ggttactgga acgttgtgag ggtaaacaac tggcggtatg gatgcggcgg gaccagagaa
     3001 aaatcactca gggtcaatgc cagcgcttcg ttaatacaga tgtaggtgtt ccacagggta
     3061 gccagcagca tcctgcgatg cagatccgga acataatggt gcagggcgct gacttccgcg
     3121 tttccagact ttacgaaaca cggaaaccga agaccattca tgttgttgct caggtcgcag
     3181 acgttttgca gcagcagtcg cttcacgttc gctcgcgtat cggtgattca ttctgctaac
     3241 cagtaaggca accccgccag cctagccggg tcctcaacga caggagcacg atcatgcgca
     3301 cccgtggggc cgccatgccg gcgataatgg cctgcttctc gccgaaacgt ttggtggcgg
     3361 gaccagtgac gaaggcttga gcgagggcgt gcaagattcc gaataccgca agcgacaggc
     3421 cgatcatcgt cgcgctccag cgaaagcggt cctcgccgaa aatgacccag agcgctgccg
     3481 gcacctgtcc tacgagttgc atgataaaga agacagtcat aagtgcggcg acgatagtca
     3541 tgccccgcgc ccaccggaag gagctgactg ggttgaaggc tctcaagggc atcggtcgag
     3601 atcccggtgc ctaatgagtg agctaactta cattaattgc gttgcgctca ctgcccgctt
     3661 tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag
     3721 gcggtttgcg tattgggcgc cagggtggtt tttcttttca ccagtgagac gggcaacagc
     3781 tgattgccct tcaccgcctg gccctgagag agttgcagca agcggtccac gctggtttgc
     3841 cccagcaggc gaaaatcctg tttgatggtg gttaacggcg ggatataaca tgagctgtct
     3901 tcggtatcgt cgtatcccac taccgagata tccgcaccaa cgcgcagccc ggactcggta
     3961 atggcgcgca ttgcgcccag cgccatctga tcgttggcaa ccagcatcgc agtgggaacg
     4021 atgccctcat tcagcatttg catggtttgt tgaaaaccgg acatggcact ccagtcgcct
     4081 tcccgttccg ctatcggctg aatttgattg cgagtgagat atttatgcca gccagccaga
     4141 cgcagacgcg ccgagacaga acttaatggg cccgctaaca gcgcgatttg ctggtgaccc
     4201 aatgcgacca gatgctccac gcccagtcgc gtaccgtctt catgggagaa aataatactg
     4261 ttgatgggtg tctggtcaga gacatcaaga aataacgccg gaacattagt gcaggcagct
     4321 tccacagcaa tggcatcctg gtcatccagc ggatagttaa tgatcagccc actgacgcgt
     4381 tgcgcgagaa gattgtgcac cgccgcttta caggcttcga cgccgcttcg ttctaccatc
     4441 gacaccacca cgctggcacc cagttgatcg gcgcgagatt taatcgccgc gacaatttgc
     4501 gacggcgcgt gcagggccag actggaggtg gcaacgccaa tcagcaacga ctgtttgccc
     4561 gccagttgtt gtgccacgcg gttgggaatg taattcagct ccgccatcgc cgcttccact
     4621 ttttcccgcg ttttcgcaga aacgtggctg gcctggttca ccacgcggga aacggtctga
     4681 taagagacac cggcatactc tgcgacatcg tataacgtta ctggtttcac attcaccacc
     4741 ctgaattgac tctcttccgg gcgctatcat gccataccgc gaaaggtttt gcgccattcg
     4801 atggtgtccg ggatctcgac gctctccctt atgcgactcc tgcattagga agcagcccag
     4861 tagtaggttg aggccgttga gcaccgccgc cgcaaggaat ggtgcatgca aggagatggc
     4921 gcccaacagt cccccggcca cggggcctgc caccataccc acgccgaaac aagcgctcat
     4981 gagcccgaag tggcgagccc gatcttcccc atcggtgatg tcggcgatat aggcgccagc
     5041 aaccgcacct gtggcgccgg tgatgccggc cacgatgcgt ccggcgtaga ggatcgagat
     5101 ctcgatcccg cgaaattaat acgactcact ataggggaat tgtgagcgga taacaattcc
     5161 cctctagaaa taattttgtt taactttaag aaggagatat acatatgcag accaccgtga
     5221 ccaccatgct cctcgcgtcc gtagcattag cggcttgcgc cggaggaggt tccactcctc
     5281 tgcctctacc gcagcagcag ccgcctcagc aggaaccgcc acctcctcct gttccgctag
     5341 ccagtcgcgc cgcgtgtgag gcgctcaaag atggtaatgg cgacatggtt tggccgaatg
     5401 ccgccacggt tgtagaggtt gcagcctggc gtgatgcagc accggccacg gcatcagccg
     5461 cagccctgcc ggagcattgc gaagtatcag gcgcgattgc caagcgtact gggattgatg
     5521 ggtacccgta tgaaattaag tttcgcctgc gcatgcccgc tgagtggaac ggccgttttt
     5581 tcatggaggg tggcagtggt acgaacggct ctctctcagc ggcgaccgga agtatcggcg
     5641 gcggtcagat cgcctcagcg ctgagtcgta actttgcaac aattgctacc gacggaggac
     5701 atgacaatgc ggtgaatgat aatccggatg cgctcggtac cgtcgcattt ggtctcgatc
     5761 cccaggcacg cttagacatg ggctacaact cctatgatca ggtgactcag gccggcaaag
     5821 ccgccgttgc acgcttttat ggtcgcgcag ccgacaagag ctacttcatc ggctgttcgg
     5881 agggcggccg cgagggcatg atgctgtccc agcgctttcc atcacattac gatggcattg
     5941 tggcgggcgc accgggatat cagttgccga aggccggaat tagtggcgcg tggaccaccc
     6001 agagcttagc gcccgccgcc gttggcctgg atgcccaggg agtgccgctg attaataaga
     6061 gcttttctga cgcagacctc catttactgt cgcaggcgat tctcggaaca tgcgacgcct
     6121 tggatggcct ggccgacggc atcgttgaca actaccgagc gtgccaagcg gcttttgatc
     6181 cggcgactgc agccaaccca gcgaatggcc aagccctgca gtgcgtgggc gcaaagacag
     6241 ccgattgctt atcgcccgtc caagttacgg cgattaaacg agcgatggcc ggtccggtaa
     6301 atagcgcggg tacgccgtta tataatagat gggcctggga cgcaggtatg agcggtctta
     6361 gtggtaccac ttacaatcag ggttggcgca gctggtggct gggatcgttt aacagctcgg
     6421 cgaataacgc acaacgtgta tctggtttct cagcgcggag ctggctggtg gactttgcta
     6481 ccccgccgga gccgatgccc atgacccaag tcgccgcccg tatgatgaaa tttgatttcg
     6541 atatcgatcc tctgaaaata tgggctactt cgggccaatt tacccagagt agtatggact
     6601 ggcacggtgc cactagcacc gaccttgctg cctttcggga ccgcggcggt aaaatgattc
     6661 tgtatcactg catgagcgat gccgcattct ctgcactaga tacagcagat tattatgaac
     6721 gcctgggtgc cgcaatgccg ggcgccgcgg gctttgctcg tctgttcttg gttccgggaa
     6781 tgaaccattg ctgcgggggt ccaggtaccg accgctttga tatgctaaca ccgttagttg
     6841 catgggttga acgtggggaa gcccctgacc aaattagcgc ctggagcggc acccccggct
     6901 actttggtgt ggccgcccgc actcgaccgt tatgtcccta tccgcagatt gcgcgctata
     6961 agggatcagg cgatatcaat accgaagcaa attttgcgtg tgccgctcca ccgctcgagc
     7021 accaccatca ccaccactga gatccggctg ctaacaaagc ccgaaaggaa gctgagttgg
     7081 ctgctgccac cgctgagcaa taactagcat aaccccttgg ggcctctaaa cgggtcttga
     7141 ggggtttttt gctgaaagga ggaactatat ccggat
//
