LOCUS       Exported                6680 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    pcDNA3-EGFP-RhoA-wt
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6680)
  AUTHORS   Subauste MC, Von Herrath M, Benard V, Chamberlain CE, Chuang TH, Chu
            K, Bokoch GM, Hahn KM
  TITLE     Rho family proteins modulate rapid apoptosis induced by cytotoxic T 
            lymphocytes and Fas.
  JOURNAL   J Biol Chem. 2000 Mar 31. 275(13):9725-33.
  PUBMED    10734125
REFERENCE   2  (bases 1 to 6680)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6680
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     primer_bind     complement(46..63)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(217..805)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(297..316)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(976..1836)
                     /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     1599..1618
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1837..1941)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(2016..2035)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        2207..2586
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2587..2790
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2741..2761
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     2835..2854
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2835..2853
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      2867..2876
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             2873..3589
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             2873..3589
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2918..2939)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3179..3198)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3526..3547
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             3596..4177
                     /codon_start=1
                     /product="human Rho family small GTPase RhoA"
                     /label=RhoA
                     /translation="MAAIRKKLVIVGDGACGKTCLLIVFSKDQFPEVYVPTVFENYVAD
                     IEVDGKQVELALWDTAGQEDYDRLRPLSYPDTDVILMCFSIDSPDSLENIPEKWTPEVK
                     HFCPNVPIILVGNKKDLRNDEHTRRELAKMKQEPVKPEEGRDMANRIGAFGYMECSAKT
                     KDGVREVFEMATRAALQARRGKKKSGCLVL"
     promoter        complement(4205..4223)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(4206..4223)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(4243..4260)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4249..4473
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      4519..4947
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4606..4625)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4816..4837
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4956..4976)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4961..5290
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5141..5276
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     5203..5222
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             5357..6151
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(5411..5430)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     6021..6040
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    6325..6446
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6362..6381)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6416..6435
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(6495..6511)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6495..6511)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6508..6530)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6519..6535
                     /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)."
     promoter        complement(6543..6573)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6588..6609
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
ORIGIN
        1 acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc
       61 gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg
      121 gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa
      181 ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga
      241 cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag
      301 ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct
      361 taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg
      421 ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc
      481 ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt
      541 aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta
      601 tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac
      661 agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc
      721 ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat
      781 tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc
      841 tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt
      901 cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta
      961 aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct
     1021 atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg
     1081 cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga
     1141 tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt
     1201 atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt
     1261 taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt
     1321 tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat
     1381 gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc
     1441 cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc
     1501 cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat
     1561 gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag
     1621 aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt
     1681 accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc
     1741 ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa
     1801 gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg
     1861 aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa
     1921 taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg tcgacggatc
     1981 gggagatctc ccgatcccct atggtgcact ctcagtacaa tctgctctga tgccgcatag
     2041 ttaagccagt atctgctccc tgcttgtgtg ttggaggtcg ctgagtagtg cgcgagcaaa
     2101 atttaagcta caacaaggca aggcttgacc gacaattgca tgaagaatct gcttagggtt
     2161 aggcgttttg cgctgcttcg cgatgtacgg gccagatata cgcgttgaca ttgattattg
     2221 actagttatt aatagtaatc aattacgggg tcattagttc atagcccata tatggagttc
     2281 cgcgttacat aacttacggt aaatggcccg cctggctgac cgcccaacga cccccgccca
     2341 ttgacgtcaa taatgacgta tgttcccata gtaacgccaa tagggacttt ccattgacgt
     2401 caatgggtgg agtatttacg gtaaactgcc cacttggcag tacatcaagt gtatcatatg
     2461 ccaagtacgc cccctattga cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag
     2521 tacatgacct tatgggactt tcctacttgg cagtacatct acgtattagt catcgctatt
     2581 accatggtga tgcggttttg gcagtacatc aatgggcgtg gatagcggtt tgactcacgg
     2641 ggatttccaa gtctccaccc cattgacgtc aatgggagtt tgttttggca ccaaaatcaa
     2701 cgggactttc caaaatgtcg taacaactcc gccccattga cgcaaatggg cggtaggcgt
     2761 gtacggtggg aggtctatat aagcagagct ctctggctaa ctagagaacc cactgcttac
     2821 tggcttatcg aaattaatac gactcactat agggagaccc aagcttgcca ccatggtgag
     2881 caagggcgag gagctgttca ccggggtggt gcccatcctg gtcgagctgg acggcgacgt
     2941 aaacggccac aagttcagcg tgtccggcga gggcgagggc gatgccacct acggcaagct
     3001 gaccctgaag ttcatctgca ccaccggcaa gctgcccgtg ccctggccca ccctcgtgac
     3061 caccctgacc tacggcgtgc agtgcttcag ccgctacccc gaccacatga agcagcacga
     3121 cttcttcaag tccgccatgc ccgaaggcta cgtccaggag cgcaccatct tcttcaagga
     3181 cgacggcaac tacaagaccc gcgccgaggt gaagttcgag ggcgacaccc tggtgaaccg
     3241 catcgagctg aagggcatcg acttcaagga ggacggcaac atcctggggc acaagctgga
     3301 gtacaactac aacagccaca acgtctatat catggccgac aagcagaaga acggcatcaa
     3361 ggtgaacttc aagatccgcc acaacatcga ggacggcagc gtgcagctcg ccgaccacta
     3421 ccagcagaac acccccatcg gcgacggccc cgtgctgctg cccgacaacc actacctgag
     3481 cacccagtcc gccctgagca aagaccccaa cgagaagcgc gatcacatgg tcctgctgga
     3541 gttcgtgacc gccgccggga tcactctcgg catggacgag ctgtacaagg aattcatggc
     3601 tgccatccgg aagaaactgg tgattgttgg tgatggagcc tgtggaaaga catgcttgct
     3661 catagtcttc agcaaggacc agttcccaga ggtgtatgtg cccacagtgt ttgagaacta
     3721 tgtggcagat atcgaggtgg atggaaagca ggtagagttg gctttgtggg acacagctgg
     3781 gcaggaagat tatgatcgcc tgaggcccct ctcctaccca gataccgatg ttatactgat
     3841 gtgtttttcc atcgacagcc ctgatagttt agaaaacatc ccagaaaagt ggaccccaga
     3901 agtcaagcat ttctgtccca acgtgcccat catcctggtt gggaataaga aggatcttcg
     3961 gaatgatgag cacacaaggc gggagctagc caagatgaag caggagccgg tgaaacctga
     4021 agaaggcaga gatatggcaa acaggattgg cgcttttggg tacatggagt gttcagcaaa
     4081 gaccaaagat ggagtgagag aggtttttga aatggctacg agagctgctc tgcaagctag
     4141 acgtgggaag aaaaaatctg gttgccttgt cttgtgactc gagcatgcat ctagagggcc
     4201 ctattctata gtgtcaccta aatgctagag ctcgctgatc agcctcgact gtgccttcta
     4261 gttgccagcc atctgttgtt tgcccctccc ccgtgccttc cttgaccctg gaaggtgcca
     4321 ctcccactgt cctttcctaa taaaatgagg aaattgcatc gcattgtctg agtaggtgtc
     4381 attctattct ggggggtggg gtggggcagg acagcaaggg ggaggattgg gaagacaata
     4441 gcaggcatgc tggggatgcg gtgggctcta tggcttctga ggcggaaaga accagctggg
     4501 gctctagggg gtatccccac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg
     4561 ttacgcgcag cgtgaccgct acacttgcca gcgccctagc gcccgctcct ttcgctttct
     4621 tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat cgggggctcc
     4681 ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt gattagggtg
     4741 atggttcacg tagtgggcca tcgccctgat agacggtttt tcgccctttg acgttggagt
     4801 ccacgttctt taatagtgga ctcttgttcc aaactggaac aacactcaac cctatctcgg
     4861 tctattcttt tgatttataa gggattttgc cgatttcggc ctattggtta aaaaatgagc
     4921 tgatttaaca aaaatttaac gcgaattaat tctgtggaat gtgtgtcagt tagggtgtgg
     4981 aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc
     5041 aaccaggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct
     5101 caattagtca gcaaccatag tcccgcccct aactccgccc atcccgcccc taactccgcc
     5161 cagttccgcc cattctccgc cccatggctg actaattttt tttatttatg cagaggccga
     5221 ggccgcctct gcctctgagc tattccagaa gtagtgagga ggcttttttg gaggcctagg
     5281 cttttgcaaa aagctcccgg gagcttgtat atccattttc ggatctgatc aagagacagg
     5341 atgaggatcg tttcgcatga ttgaacaaga tggattgcac gcaggttctc cggccgcttg
     5401 ggtggagagg ctattcggct atgactgggc acaacagaca atcggctgct ctgatgccgc
     5461 cgtgttccgg ctgtcagcgc aggggcgccc ggttcttttt gtcaagaccg acctgtccgg
     5521 tgccctgaat gaactgcagg acgaggcagc gcggctatcg tggctggcca cgacgggcgt
     5581 tccttgcgca gctgtgctcg acgttgtcac tgaagcggga agggactggc tgctattggg
     5641 cgaagtgccg gggcaggatc tcctgtcatc tcaccttgct cctgccgaga aagtatccat
     5701 catggctgat gcaatgcggc ggctgcatac gcttgatccg gctacctgcc cattcgacca
     5761 ccaagcgaaa catcgcatcg agcgagcacg tactcggatg gaagccggtc ttgtcgatca
     5821 ggatgatctg gacgaagagc atcaggggct cgcgccagcc gaactgttcg ccaggctcaa
     5881 ggcgcgcatg cccgacggcg aggatctcgt cgtgacccat ggcgatgcct gcttgccgaa
     5941 tatcatggtg gaaaatggcc gcttttctgg attcatcgac tgtggccggc tgggtgtggc
     6001 ggaccgctat caggacatag cgttggctac ccgtgatatt gctgaagagc ttggcggcga
     6061 atgggctgac cgcttcctcg tgctttacgg tatcgccgct cccgattcgc agcgcatcgc
     6121 cttctatcgc cttcttgacg agttcttctg agcgggactc tggggttcga aatgaccgac
     6181 caagcgacgc ccaacctgcc atcacgagat ttcgattcca ccgccgcctt ctatgaaagg
     6241 ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga tcctccagcg cggggatctc
     6301 atgctggagt tcttcgccca ccccaacttg tttattgcag cttataatgg ttacaaataa
     6361 agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt
     6421 ttgtccaaac tcatcaatgt atcttatcat gtctgtatac cgtcgacctc tagctagagc
     6481 ttggcgtaat catggtcata gctgtttcct gtgtgaaatt gttatccgct cacaattcca
     6541 cacaacatac gagccggaag cataaagtgt aaagcctggg gtgcctaatg agtgagctaa
     6601 ctcacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag
     6661 ctgcattaat gaatcggcca
//
