LOCUS       Exported                5862 bp ds-DNA     circular SYN 12-MAR-2019
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  recombinant plasmid
REFERENCE   1  (bases 1 to 5862)
  AUTHORS   Jason Vevea
  TITLE     Direct Submission
  JOURNAL   Exported Thursday, Jul 23, 2020 from SnapGene Viewer 5.0.8
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..5862
                     /organism="recombinant plasmid"
                     /mol_type="other DNA"
     promoter        36..1217
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          266..1208
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     misc_feature    1258..1884
                     /label=HCV NS3/4a FAST protease
                     /note="NS3/4a protease FAST VARIANT. Sequence from 
                     PSD95-TS2-YFP.
                     
                     
                     Butko, M.T., Yang, J., Geng, Y., Kim, H.J., Jeon, N.L., 
                     Shu, X., Mackey, M.R., Ellisman, M.H., Tsien, R.Y. and Lin,
                     M.Z., 2012. Fluorescent and photo-oxidizing TimeSTAMP tags 
                     track protein fates in light and electron microscopy. 
                     Nature neuroscience, 15(12), pp.1742-1751.
                     "
     misc_feature    1885..1926
                     /label=GS(GSS)4 Linker 3
     CDS             1927..2622
                     /codon_start=1
                     /label=msGFP
                     /translation="MDSTESLFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTITFKDDG
                     TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA
                     NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLLE
                     FVTAAGITHG"
     primer_bind     1927..1962
                     /label=DSTES
                     /note="Use together with BL14 to amplify msGFP, msYFP, and 
                     msCFP with the improved N-terminal codons from A24. 
                     Subclone the PCR product back into the same vector using 
                     BamHI and NotI."
     primer_bind     1930..1953
                     /label=new N GFP lin F
     primer_bind     complement(2605..2630)
                     /label=BL14
                     /note="This oligo (used with BL13) will replace the linker 
                     MDELYK to GSSGSS in monomeric superfolder GFP, CFP and 
                     YFP."
     primer_bind     complement(2612..2630)
                     /label=BL24
                     /note="This oligo is used to amplify msfFPs (Green, Cyan, 
                     Yellow) from the pMSGFP vector into the NT-HT vector for 
                     the Tub1 fussion.  The cut sites are BspEI for the 
                     N-terminal and SacI for the C-terminal. Use this oligo with
                     BL23."
     misc_feature    2623..2664
                     /label=GS(GSS)4 Linker 1
     CDS             2701..2802
                     /codon_start=1
                     /product="Modified OMP25 c terminal targeting sequencecyto 
                     aa = GSKRG
                     "
                     /label=OMP25 v2 mito targeting sequence
                     /translation="GSKRGVPVAVVLLPVFALTLVAVWAFVRYRKQL"
     misc_feature    2716..2778
                     /label=OMP25 v2 TMD
     polyA_site      2951..3006
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal"
     primer_bind     complement(3365..3381)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    3389..3405
                     /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(3413..3443)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3458..3479
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        3538..3733
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      3584..3719
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    3739..3873
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     rep_origin      complement(4112..4700)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(4871..5731)
                     /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"
     promoter        complement(5732..5836)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gtcgacattg attattgact agatcatcgc gtgaggctcc ggtgcccgtc agtgggcaga
       61 gcgcacatcg cccacagtcc ccgagaagtt ggggggaggg gtcggcaatt gaaccggtgc
      121 ctagagaagg tggcgcgggg taaactggga aagtgatgtc gtgtactggc tccgcctttt
      181 tcccgagggt gggggagaac cgtatataag tgcagtagtc gccgtgaacg ttctttttcg
      241 caacgggttt gccgccagaa cacaggtaag tgccgtgtgt ggttcccgcg ggcctggcct
      301 ctttacgggt tatggccctt gcgtgccttg aattacttcc acgcccctgg ctgcagtacg
      361 tgattcttga tcccgagctt cgggttggaa gtgggtggga gagttcgagg ccttgcgctt
      421 aaggagcccc ttcgcctcgt gcttgagttg aggcctggcc tgggcgctgg ggccgccgcg
      481 tgcgaatctg gtggcacctt cgcgcctgtc tcgctgcttt cgataagtct ctagccattt
      541 aaaatttttg atgacctgct gcgacgcttt ttttctggca agatagtctt gtaaatgcgg
      601 gccaagatct gcacactggt atttcggttt ttggggccgc gggcggcgac ggggcccgtg
      661 cgtcccagcg cacatgttcg gcgaggcggg gcctgcgagc gcggccaccg agaatcggac
      721 gggggtagtc tcaagctggc cggcctgctc tggtgcctgg cctcgcgccg ccgtgtatcg
      781 ccccgccctg ggcggcaagg ctggcccggt cggcaccagt tgcgtgagcg gaaagatggc
      841 cgcttcccgg ccctgctgca gggagctcaa aatggaggac gcggcgctcg ggagagcggg
      901 cgggtgagtc acccacacaa aggaaaaggg cctttccgtc ctcagccgtc gcttcatgtg
      961 actccacgga gtaccgggcg ccgtccaggc acctcgatta gttctcgagc ttttggagta
     1021 cgtcgtcttt aggttggggg gaggggtttt atgcgatgga gtttccccac actgagtggg
     1081 tggagactga agttaggcca gcttggcact tgatgtaatt ctccttggaa tttgcccttt
     1141 ttgagtttgg atcttggttc attctcaagc ctcagacagt ggttcaaagt ttttttcttc
     1201 catttcaggt gtcgtgagga attctgcagt cgacggtacc gcgggcccgc caccatgggc
     1261 tgcgtggtga tcgtgggcag gatcgtgctg agcggcagcg gcaccagcgc gcccatcacg
     1321 gcgtacgccc agcagacgag aggcctccta gggtgtataa tcaccagcct gactggccgg
     1381 gacaaaaacc aagtggaggg tgaggtccag atcgtgtcaa ctgctaccca aaccttcctg
     1441 gcaacgtgca tcaatggggt atgctggaca gtctaccacg gggccggaac gaggaccatc
     1501 gcatcaccca agggtcctgt catccagatg tataccaatg tggaccaaga ccttgtgggc
     1561 tggcccgctc ctcaaggttc ccgctcattg acaccctgta cctgcggctc ctcggacctt
     1621 tacctggtca cgaggcacgc cgatgtcatt cccgtgcgcc ggcgaggtga tagcaggggt
     1681 agcctgcttt cgccccggcc catttcctac ttgaaaggct cctcgggggg tccgctgttg
     1741 tgccccgcgg gacacgccgt gggcctattc agggccgcgg tgtgcacccg tggagtggct
     1801 aaagcggtgg actttatccc tgtggagaac ctagagacaa ccatgagatc cccggtgttc
     1861 acggacaact cctctccacc agcagggtca ggttcttcgg gcagttcggg gtccagcgga
     1921 tctagtatgg atagcactga gagcctgttc accggggtgg tgcccatcct ggtcgagctg
     1981 gacggcgacg taaacggcca caagttcagc gtgcgcggcg agggcgaggg cgatgccacc
     2041 aacggcaagc tgaccctgaa gttcatctgc accaccggca agctgcccgt gccctggccc
     2101 accctcgtga ccaccctgac ctacggcgtg cagtgcttca gccgctaccc cgaccacatg
     2161 aagcagcacg acttcttcaa gtccgccatg cccgaaggct acgtccagga gcgcaccatc
     2221 accttcaagg acgacggcac ctacaagacc cgcgccgagg tgaagttcga gggcgacacc
     2281 ctggtgaacc gcatcgagct gaagggcatc gacttcaagg aggacggcaa catcctgggg
     2341 cacaagctgg agtacaactt caacagccac aacgtctata tcaccgccga caagcagaag
     2401 aacggcatca aggccaactt caagatccgc cacaacgtgg aggacggcag cgtgcagctc
     2461 gccgaccact accagcagaa cacccccatc ggcgacggcc ccgtgctgct gcccgacaac
     2521 cactacctga gcacccagtc caagctgagc aaagacccca acgagaagcg cgatcacatg
     2581 gtcctgctgg agttcgtgac cgccgccggg atcactcacg gcggatccgg aagttcgggg
     2641 tccagcggtt cttcaggcag ttccgaggat gtggtctgct gtcagatgtc ctacggcagt
     2701 ggatctaagc gcggagttcc tgtagctgtg gtgctgctgc cagtgtttgc ccttaccctg
     2761 gtagcagttt gggccttcgt gagataccga aagcagctct gaggccgcac tcctcaggtg
     2821 caggctgcct atcagaaggt ggtggctggt gtggccaatg ccctggctca caaataccac
     2881 tgagatcttt ttccctctgc caaaaattat ggggacatca tgaagcccct tgagcatctg
     2941 acttctggct aataaaggaa atttattttc attgcaatag tgtgttggaa ttttttgtgt
     3001 ctctcactcg gaaggacata tgggagggca aatcatttaa aacatcagaa tgagtatttg
     3061 gtttagagtt tggcaacata tgccatatgc tggctgccat gaacaaaggt ggctataaag
     3121 aggtcatcag tatatgaaac agccccctgc tgtccattcc ttattccata gaaaagcctt
     3181 gacttgaggt tagatttttt ttatattttg ttttgtgtta tttttttctt taacatccct
     3241 aaaattttcc ttacatgttt tactagccag atttttcctc ctctcctgac tactcccagt
     3301 catagctgtc cctcttctct tatgaagatc cctcgacctg cagcccaagc ttggcgtaat
     3361 catggtcata gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacaacatac
     3421 gagccggaag cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa
     3481 ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag cggatccgca
     3541 tctcaattag tcagcaacca tagtcccgcc cctaactccg cccatcccgc ccctaactcc
     3601 gcccagttcc gcccattctc cgccccatgg ctgactaatt ttttttattt atgcagaggc
     3661 cgaggccgcc tcggcctctg agctattcca gaagtagtga ggaggctttt ttggaggcct
     3721 aggcttttgc aaaaagctaa cttgtttatt gcagcttata atggttacaa ataaagcaat
     3781 agcatcacaa atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc
     3841 aaactcatca atgtatctta tcatgtctgg atccgctgca ttaatgaatc ggccaacgcg
     3901 cggggagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact gactcgctgc
     3961 gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat
     4021 ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca
     4081 ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc
     4141 atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc
     4201 aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg
     4261 gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcaatgc tcacgctgta
     4321 ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg
     4381 ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac
     4441 acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag
     4501 gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga aggacagtat
     4561 ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat
     4621 ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc
     4681 gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt
     4741 ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct
     4801 agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt
     4861 ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc
     4921 gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac
     4981 catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat
     5041 cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg
     5101 cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata
     5161 gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta
     5221 tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt
     5281 gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag
     5341 tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa
     5401 gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc
     5461 gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt
     5521 taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc
     5581 tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta
     5641 ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa
     5701 taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca
     5761 tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac
     5821 aaataggggt tccgcgcaca tttccccgaa aagtgccacc tg
//
