LOCUS       Exported                6047 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  express NS2B-NS3 protease in E.coli.
ACCESSION   .
VERSION     .
KEYWORDS    bZiPro
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6047)
  AUTHORS   Zhang Z, Li Y, Loh YR, Phoo WW, Hung AW, Kang C, Luo D
  TITLE     Crystal structure of unlinked NS2B-NS3 protease from Zika virus.
  JOURNAL   Science. 2016 Dec 8. pii: aai9309.
  PUBMED    27940580
REFERENCE   2  (bases 1 to 6047)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6047
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     protein_bind    3..27
                     /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)."
     RBS             42..64
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             83..100
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             110..127
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     promoter        733..751
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    752..776
                     /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)."
     CDS             996..1040
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     primer_bind     complement(1078..1096)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1092..1139
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1176..1631
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1263..1282)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1473..1494
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1746..2606)
                     /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     2369..2388
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2607..2699)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      2780..3368
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3269..3288
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3522..3539
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3554..3693
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3619..3638)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3779..3801
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3795..3986)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4558..5640)
                     /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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     5602..5621
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5641..5718)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     5924..5943
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        6031..2
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 ggggaattgt gagcggataa caattcccct ctagaaataa ttttgtttaa ctttaagaag
       61 gagatatacc atgggcagca gccatcatca tcatcatcat agcagcggcc tggtgccgcg
      121 cggcagcggt gccctctggg acgtgcctgc tcctaaagaa gtgaagaagg gagagactac
      181 agatggagtg taccgtgtta tgactcgccg cctgctgggt tcaacacagg ttggagtggg
      241 agtcatgcaa gagggagtct tccataccat gtggcacgtc acgaaaggag ccgcattgcg
      301 tagcggtgaa ggacgccttg atccatactg gggagacgtc aagcaggacc tggtgtcata
      361 ttgtgggccg tggaagttgg atgcagcctg ggatggactc agtgaggtgc agctcttggc
      421 cgtacctcct ggagagcgtg ctaagaacat tcagactctg cctggaatct ttaagaccaa
      481 ggatggtgac atcggagcag ttgctctcga ctaccctgca ggaacctcag gatctccgat
      541 cctcgacaag tgcggacgtg tgatcggact ttatggcaat ggtgttgtga tcaagaatgg
      601 aagctatgtt agtgccataa cccagggcaa gcgtgaggag gagactccgg ttgagtgatg
      661 aaagcttgcg gccgcataat gcttaagtcg aacagaaagt aatcgtattg tacacggccg
      721 cataatcgaa attaatacga ctcactatag gggaattgtg agcggataac aattccccat
      781 cttagtatat tagttaagta taagaaggag atatacatat gacgggaaag agtgtggaca
      841 tgtacattga acgtgcaggt gatattacat gggagaaaga cgcggaagtc actggaaaca
      901 gtcctcgtct tgacgtggca cttgatgaga gtggtgattt ctctctggta gaggaggatg
      961 gaccacctat gcgtgagtaa tgactcgagt ctggtaaaga aaccgctgct gcgaaatttg
     1021 aacgccagca catggactcg tctactagcg cagcttaatt aacctaggct gctgccaccg
     1081 ctgagcaata actagcataa ccccttgggg cctctaaacg ggtcttgagg ggttttttgc
     1141 tgaaaggagg aactatatcc ggattggcga atgggacgcg ccctgtagcg gcgcattaag
     1201 cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc
     1261 cgctcctttc gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc
     1321 tctaaatcgg gggctccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa
     1381 aaaacttgat tagggtgatg gttcacgtag tgggccatcg ccctgataga cggtttttcg
     1441 ccctttgacg ttggagtcca cgttctttaa tagtggactc ttgttccaaa ctggaacaac
     1501 actcaaccct atctcggtct attcttttga tttataaggg attttgccga tttcggccta
     1561 ttggttaaaa aatgagctga tttaacaaaa atttaacgcg aattttaaca aaatattaac
     1621 gtttacaatt tctggcggca cgatggcatg agattatcaa aaaggatctt cacctagatc
     1681 cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct
     1741 gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca
     1801 tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct
     1861 ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca
     1921 ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc
     1981 atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg
     2041 cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct
     2101 tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa
     2161 aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta
     2221 tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc
     2281 ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg
     2341 agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa
     2401 gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg
     2461 agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc
     2521 accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg
     2581 gcgacacgga aatgttgaat actcatactc ttcctttttc aatcatgatt gaagcattta
     2641 tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat
     2701 aggtcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag
     2761 aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa
     2821 caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt
     2881 ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgtcctt ctagtgtagc
     2941 cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa
     3001 tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa
     3061 gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc
     3121 ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa
     3181 gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa
     3241 caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg
     3301 ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc
     3361 tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg
     3421 ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg
     3481 agtgagctga taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg
     3541 aagcggaaga gcgcctgatg cggtattttc tccttacgca tctgtgcggt atttcacacc
     3601 gcaatggtgc actctcagta caatctgctc tgatgccgca tagttaagcc agtatacact
     3661 ccgctatcgc tacgtgactg ggtcatggct gcgccccgac acccgccaac acccgctgac
     3721 gcgccctgac gggcttgtct gctcccggca tccgcttaca gacaagctgt gaccgtctcc
     3781 gggagctgca tgtgtcagag gttttcaccg tcatcaccga aacgcgcgag gcagctgcgg
     3841 taaagctcat cagcgtggtc gtgaagcgat tcacagatgt ctgcctgttc atccgcgtcc
     3901 agctcgttga gtttctccag aagcgttaat gtctggcttc tgataaagcg ggccatgtta
     3961 agggcggttt tttcctgttt ggtcactgat gcctccgtgt aagggggatt tctgttcatg
     4021 ggggtaatga taccgatgaa acgagagagg atgctcacga tacgggttac tgatgatgaa
     4081 catgcccggt tactggaacg ttgtgagggt aaacaactgg cggtatggat gcggcgggac
     4141 cagagaaaaa tcactcaggg tcaatgccag cgcttcgtta atacagatgt aggtgttcca
     4201 cagggtagcc agcagcatcc tgcgatgcag atccggaaca taatggtgca gggcgctgac
     4261 ttccgcgttt ccagacttta cgaaacacgg aaaccgaaga ccattcatgt tgttgctcag
     4321 gtcgcagacg ttttgcagca gcagtcgctt cacgttcgct cgcgtatcgg tgattcattc
     4381 tgctaaccag taaggcaacc ccgccagcct agccgggtcc tcaacgacag gagcacgatc
     4441 atgctagtca tgccccgcgc ccaccggaag gagctgactg ggttgaaggc tctcaagggc
     4501 atcggtcgag atcccggtgc ctaatgagtg agctaactta cattaattgc gttgcgctca
     4561 ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc
     4621 gcggggagag gcggtttgcg tattgggcgc cagggtggtt tttcttttca ccagtgagac
     4681 gggcaacagc tgattgccct tcaccgcctg gccctgagag agttgcagca agcggtccac
     4741 gctggtttgc cccagcaggc gaaaatcctg tttgatggtg gttaacggcg ggatataaca
     4801 tgagctgtct tcggtatcgt cgtatcccac taccgagatg tccgcaccaa cgcgcagccc
     4861 ggactcggta atggcgcgca ttgcgcccag cgccatctga tcgttggcaa ccagcatcgc
     4921 agtgggaacg atgccctcat tcagcatttg catggtttgt tgaaaaccgg acatggcact
     4981 ccagtcgcct tcccgttccg ctatcggctg aatttgattg cgagtgagat atttatgcca
     5041 gccagccaga cgcagacgcg ccgagacaga acttaatggg cccgctaaca gcgcgatttg
     5101 ctggtgaccc aatgcgacca gatgctccac gcccagtcgc gtaccgtctt catgggagaa
     5161 aataatactg ttgatgggtg tctggtcaga gacatcaaga aataacgccg gaacattagt
     5221 gcaggcagct tccacagcaa tggcatcctg gtcatccagc ggatagttaa tgatcagccc
     5281 actgacgcgt tgcgcgagaa gattgtgcac cgccgcttta caggcttcga cgccgcttcg
     5341 ttctaccatc gacaccacca cgctggcacc cagttgatcg gcgcgagatt taatcgccgc
     5401 gacaatttgc gacggcgcgt gcagggccag actggaggtg gcaacgccaa tcagcaacga
     5461 ctgtttgccc gccagttgtt gtgccacgcg gttgggaatg taattcagct ccgccatcgc
     5521 cgcttccact ttttcccgcg ttttcgcaga aacgtggctg gcctggttca ccacgcggga
     5581 aacggtctga taagagacac cggcatactc tgcgacatcg tataacgtta ctggtttcac
     5641 attcaccacc ctgaattgac tctcttccgg gcgctatcat gccataccgc gaaaggtttt
     5701 gcgccattcg atggtgtccg ggatctcgac gctctccctt atgcgactcc tgcattagga
     5761 agcagcccag tagtaggttg aggccgttga gcaccgccgc cgcaaggaat ggtgcatgca
     5821 aggagatggc gcccaacagt cccccggcca cggggcctgc caccataccc acgccgaaac
     5881 aagcgctcat gagcccgaag tggcgagccc gatcttcccc atcggtgatg tcggcgatat
     5941 aggcgccagc aaccgcacct gtggcgccgg tgatgccggc cacgatgcgt ccggcgtaga
     6001 ggatcgagat cgatctcgat cccgcgaaat taatacgact cactata
//
