LOCUS       40924_44242        6745 bp DNA     circular SYN 13-MAY-2021
DEFINITION  synthetic circular DNA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6745)
  AUTHORS   Wu YI, Frey D, Lungu OI, Jaehrig A, Schlichting I, Kuhlman B, Hahn 
            KM
  TITLE     A genetically encoded photoactivatable Rac controls the motility of 
            living cells.
  JOURNAL   Nature. 2009 Sep 3. 461(7260):104-8.
  PUBMED    19693014
REFERENCE   2  (bases 1 to 6745)
  TITLE     Direct Submission
REFERENCE   3  (bases 1 to 6745)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nature. 
            2009 Sep 3. 461(7260):104-8."
COMMENT     SGRef: number: 2; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..6745
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        25..328
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        330..529
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early
                     promoter"
     primer_bind     735..757
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     819..838
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids,
                     forward primer"
     promoter        880..898
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    899..923
                     /label=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)."
     promoter        939..1048
                     /label=p10 promoter
                     /note="baculovirus promoter for expression in insect cells"
     CDS             1069..1086
                     /codon_start=1
                     /label=6xHis
                     /note="6xHis affinity tag"
                     /translation="HHHHHH"
     CDS             1093..1797
                     /codon_start=1
                     /label=mCherry
                     /note="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /translation="VSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEGT
                     QTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFE
                     DGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALKG
                     EIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERAE
                     GRHSTGGMDELYK"
     CDS             2227..2799
                     /codon_start=1
                     /label=Rac1 (T17N)
                     /note="dominant negative mutant of human Rac family small
                     GTPase 1"
                     /translation="ELIKCVVVGDGAVGKNCLLISYTTNAFPGEYIPTVFDNYSANVMV
                     DGKPVNLGLWDTAGQEDYDRLRPLSYPQTDVFLICFSLVSPASFENVRAKWYPEVRHHC
                     PNTPIILVGTKLDLRDDKDTIEKLKEKKLTPITYPQGLAMAKEIGAVKYLECSALTQRG
                     LKTVFDEAIRAVLCPPPVKKRKRKCLLL"
     CDS             2837..2869
                     /codon_start=1
                     /label=HSV tag
                     /note="HSV (herpes simplex virus) epitope tag"
                     /translation="QPELAPEDPED"
     CDS             2876..2899
                     /codon_start=1
                     /label=8xHis
                     /note="8xHis affinity tag"
                     /translation="HHHHHHHH"
     primer_bind     complement(2989..3008)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    3054..3109
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(3108..3127)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also
                     called rb-glob-pA-term-R"
     terminator      3197..3244
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     misc_recomb     3257..3962
                     /label=baculovirus recombination region (ORF1629)
                     /note="contains part of ORF1629"
     protein_bind    complement(3971..4004)
                     /label=loxP
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     rep_origin      complement(4172..4759)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(4775..5632)
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     misc_recomb     5875..6702
                     /label=baculovirus recombination region (lef2/ORF603)
                     /note="contains ORF603 and part of lef2"
     promoter        6713..6745
                     /label=CMV
                     /note="Human cytomegalovirus immediate early
                     enhancer/promoter"
ORIGIN
        1 tcatagccca tatatggagt tccgcgttac ataacttacg gtaaatggcc cgcctggctg
       61 accgcccaac gacccccgcc cattgacgtc aataatgacg tatgttccca tagtaacgcc
      121 aatagggact ttccattgac gtcaatgggt ggagtattta cggtaaactg cccacttggc
      181 agtacatcaa gtgtatcata tgccaagtcc gccccctatt gacgtcaatg acggtaaatg
      241 gcccgcctgg cattatgccc agtacatgac cttacgggac tttcctactt ggcagtacat
      301 ctacgtatta gtcatcgcta ttaccatgct gatgcggttt tggcagtaca ccaatgggcg
      361 tggatagcgg tttgactcac ggggatttcc aagtctccac cccattgacg tcaatgggag
      421 tttgttttgg caccaaaatc aacgggactt tccaaaatgt cgtaataacc ccgccccgtt
      481 gacgcaaatg ggcggtaggc gtgtacggtg ggaggtctat ataagcagac gtcgtttagt
      541 gaaccgtcag atcactagat gctttattgc ggtagtttat cacagttaaa ttgctaacgc
      601 cagtctcgaa cttaacgtgc agaagttggt cgtgaggcac tgggcaggta agtatcgggc
      661 cctttgtgcg gggggagcgg ctcggggctg tccgcggggg gacggctgcc ttcggggggg
      721 acggggcagg gcggggttcg gcttctggcg tgtgaccggc ggctctagag cctctgctaa
      781 ccatgttcat gccttcttct ttttcctaca gctcctgggc aacgtgctgg ttattgtgct
      841 gtctcatcat tttggcaaag aattggatcg gaccgaaatt aatacgactc actatagggg
      901 aattgtgagc ggataacaat tccccggagt taatccggga cctttaattc aacccaacac
      961 aatatattat agttaaataa gaattattat caaatcattt gtatattaat taaaatacta
     1021 tactgtaaat tacattttat ttacaatcaa aggagatata ccatggcaca ccatcaccac
     1081 catcacggat ctgtgagcaa gggcgaggag gataacatgg ccatcatcaa ggagttcatg
     1141 cgcttcaagg tgcacatgga gggctccgtg aacggccacg agttcgagat cgagggcgag
     1201 ggcgagggcc gcccctacga gggcacccag accgccaagc tgaaggtgac caagggtggc
     1261 cccctgccct tcgcctggga catcctgtcc cctcagttca tgtacggctc caaggcctac
     1321 gtgaagcacc ccgccgacat ccccgactac ttgaagctgt ccttccccga gggcttcaag
     1381 tgggagcgcg tgatgaactt cgaggacggc ggcgtggtga ccgtgaccca ggactcctcc
     1441 ctgcaggacg gcgagttcat ctacaaggtg aagctgcgcg gcaccaactt cccctccgac
     1501 ggccccgtaa tgcagaagaa gaccatgggc tgggaggcct cctccgagcg gatgtacccc
     1561 gaggacggcg ccctgaaggg cgagatcaag cagaggctga agctgaagga cggcggccac
     1621 tacgacgctg aggtcaagac cacctacaag gccaagaagc ccgtgcagct gcccggcgcc
     1681 tacaacgtca acatcaagtt ggacatcacc tcccacaacg aggactacac catcgtggaa
     1741 cagtacgaac gcgccgaggg ccgccactcc accggcggca tggacgagct gtacaaggga
     1801 tccttggcta ctacacttga acgtattgag aagaactttg tcattactga cccaagattg
     1861 ccagataatc ccattatatt cgcgtccgat agtttcttgc agttgacaga atatagccgt
     1921 gaagaaattt tgggaagaaa ctgcaggttt ctacaaggtc ctgaaactga tcgcgcgaca
     1981 gtgagaaaaa ttagagatgc catagataac caaacagagg tcactgttca gctgattaat
     2041 tatacaaaga gtggtaaaaa gttctggaac ctctttcact tgcagcctat gcgagatcag
     2101 aagggagatg tccagtactt tattggggtt cagttggatg gaactgagca tgtccgagat
     2161 gctgccgaga gagagggagt catgctgatt aagaaaactg cagaaaatat tgatgaggcg
     2221 gcaaaagaac ttatcaagtg tgtggtggtg ggagacggag ctgtaggtaa aaattgccta
     2281 ctgatcagtt acacaaccaa tgcatttcct ggagaatata tccctactgt ctttgacaat
     2341 tattctgcca atgttatggt agatggaaaa ccggtgaatc tgggcttatg ggatacagct
     2401 ggacaagaag attatgacag attacgcccc ctatcctatc cgcaaacaga tgtgttctta
     2461 atttgctttt cccttgtgag tcctgcatca tttgaaaatg tccgtgcaaa gtggtatcct
     2521 gaggtgcggc accactgtcc caacactccc atcatcctag tgggaactaa acttgatctt
     2581 agggatgata aagacacgat cgagaaactg aaggagaaga agctgactcc catcacctat
     2641 ccgcagggtc tagcaatggc taaggagatt ggtgctgtaa aatacctgga gtgctcggcg
     2701 ctcacacagc gaggcctcaa gacagtgttt gacgaagcga tccgagcagt cctctgcccg
     2761 cctcccgtga agaagaggaa gagaaaatgc ctgctgttgt aagcttgcgg ccgcacagct
     2821 gtatacacgt gcaagccagc cagaactcgc cccggaagac cccgaggatc tcgagcacca
     2881 ccatcaccat caccatcact aagtgattaa cctcaggtgc aggctgccta tcagaaggtg
     2941 gtggctggtg tggccaatgc cctggctcac aaataccact gagatcgatc tttttccctc
     3001 tgccaaaaat tatggggaca tcatgaagcc ccttgagcat ctgacttctg gctaataaag
     3061 gaaatttatt ttcattgcaa tagtgtgttg gaattttttg tgtctctcac tcggaaggac
     3121 atatgggagg gcaaatcatt taaaacatca gaatgagtat ttggtttaga gtttggcaac
     3181 atatgcccat atgtaactag cataacccct tggggcctct aaacgggtct tgaggggttt
     3241 tttgctgaaa gcatgcggag gaaattctcc ttgaagtttc cctggtgttc aaagtaaagg
     3301 agtttgcacc agacgcacct ctgttcactg gtccggcgta ttaaaacacg atacattgtt
     3361 attagtacat ttattaagcg ctagattctg tgcgttgttg atttacagac aattgttgta
     3421 cgtattttaa taattcatta aatttataat ctttagggtg gtatgttaga gcgaaaatca
     3481 aatgattttc agcgtcttta tatctgaatt taaatattaa atcctcaata gatttgtaaa
     3541 ataggtttcg attagtttca aacaagggtt gtttttccga accgatggct ggactatcta
     3601 atggattttc gctcaacgcc acaaaacttg ccaaatcttg tagcagcaat ctagctttgt
     3661 cgatattcgt ttgtgttttg ttttgtaata aaggttcgac gtcgttcaaa atattatgcg
     3721 cttttgtatt tctttcatca ctgtcgttag tgtacaattg actcgacgta aacacgttaa
     3781 atagagcttg gacatattta acatcgggcg tgttagcttt attaggccga ttatcgtcgt
     3841 cgtcccaacc ctcgtcgtta gaagttgctt ccgaagacga ttttgccata gccacacgac
     3901 gcctattaat tgtgtcggct aacacgtccg cgatcaaatt tgtagttgag ctttttggaa
     3961 ttgcgatcgc ataacttcgt atagcataca ttatacgaag ttataagctc ggaacgctgc
     4021 gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat
     4081 ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca
     4141 ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc ctgacgagca
     4201 tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca
     4261 ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg
     4321 atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag
     4381 gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt
     4441 tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca
     4501 cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg
     4561 cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa gaacagtatt
     4621 tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc
     4681 cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg
     4741 cagaaaaaaa ggatctcaag aagatccttt gttaccaatg cttaatcagt gaggcaccta
     4801 tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa
     4861 ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac
     4921 gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa
     4981 gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag
     5041 taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg
     5101 tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag
     5161 ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg
     5221 tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc
     5281 ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat
     5341 tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata
     5401 ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa
     5461 aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca
     5521 actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc
     5581 aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc
     5641 tttttcaata ttattgaagc atttatcagg gttattgtct catgtccgcg cgtttcctgc
     5701 atcttttaat caaatcccaa gatgtgtata aacgcgccgg tatgtacagg aagaggttta
     5761 tactaaactg ttacattgca aacgtggttt cgtgtgccaa gtgtgaaaac cgatgtttaa
     5821 tcaaggctct gacgcatttc tacaaccacg actccaagtg tgtgggtgaa gtcatgcatc
     5881 ttttaatcaa atcccaagat gtgtataaac caccaaactg ccaaaaaatg aaaactgtcg
     5941 acaagctctg tccgtttgct ggcaactgca agggtctcaa tcctatttgt aattattgaa
     6001 taataaaaca attataaatg tcaaatttgt tttttattaa cgatacaaac caaacgcaac
     6061 aagaacattt gtagtattat ctataattga aaacgcgtag ttataatcgc tgaggtaata
     6121 tttaaaatca ttttcaaatg attcacagtt aatttgcgac aatataattt tattttcaca
     6181 taaactagac gccttgtcgt cttcttcttc gtattccttc tctttttcat ttttctcttc
     6241 ataaaaatta acatagttat tatcgtatcc atatatgtat ctatcgtata gagtaaattt
     6301 tttgttgtca taaatatata tgtctttttt aatggggtgt atagtaccgc tgcgcatagt
     6361 ttttctgtaa tttacaacag tgctattttc tggtagttct tcggagtgtg ttgctttaat
     6421 tattaaattt atataatcaa tgaatttggg atcgtcggtt ttgtacaata tgttgccggc
     6481 atagtacgca gcttcttcta gttcaattac accatttttt agcagcaccg gattaacata
     6541 actttccaaa atgttgtacg aaccgttaaa caaaaacagt tcacctccct tttctatact
     6601 attgtctgcg agcagttgtt tgttgttaaa aataacagcc attgtaatga gacgcacaaa
     6661 ctaatatcac aaactggaaa tgtctatcaa tatatagttg ctgatggccg gcctattaat
     6721 agtaatcaat tacggggtca ttagt
//
