LOCUS       Exported                6998 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  Expresses infrared fluorescent tobacco etch virus protease sensor 
            (iTEV).
ACCESSION   .
VERSION     .
KEYWORDS    pcDNA3.1 iTEV
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6998)
  AUTHORS   To TL, Piggott BJ, Makhijani K, Yu D, Jan YN, Shu X
  TITLE     Rationally designed fluorogenic protease reporter visualizes 
            spatiotemporal dynamics of apoptosis in vivo.
  JOURNAL   Proc Natl Acad Sci U S A. 2015 Mar 17;112(11):3338-43. doi: 
            10.1073/pnas.1502857112. Epub 2015 Mar 2.
  PUBMED    25733847
REFERENCE   2  (bases 1 to 6998)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6998
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             923..1564
                     /codon_start=1
                     /product="fragment of superfolder GFP consisting of the 
                     first 10 beta-strands"
                     /label=GFP(1-10)
                     /note="associates with GFP11, the 11th beta-strand, to 
                     reconstitute fluorescent GFP (Cabantous et al., 2005; 
                     Kamiyama et al., 2016)"
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATIGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTISFKDDG
                     KYKTRAVVKFEGDTLVNRIELKGTDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA
                     NFTVRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQTVLSKDPNEK"
     CDS             2147..2167
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             2510..2557
                     /codon_start=1
                     /product="11th beta-strand of superfolder GFP"
                     /label=GFP11
                     /note="associates with GFP(1-10), consisting of the first 
                     10 beta-strands, to reconstitute fluorescent GFP (Cabantous
                     et al., 2005; Kamiyama et al., 2016)"
                     /translation="RDHMVLHEYVNAAGIT"
     primer_bind     complement(2592..2609)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2598..2822
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2868..3296
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2955..2974)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3165..3186
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3305..3325)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3310..3639
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3490..3625
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3552..3571
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3706..4500
                     /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(3760..3779)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4370..4389
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4674..4795
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4711..4730)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4765..4784
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4844..4860)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4844..4860)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4857..4879)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4868..4884
                     /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(4892..4922)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4937..4958
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5075..5092)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5246..5831)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5326..5345)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(6002..6862)
                     /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     6625..6644
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6863..6967)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc
      901 gtttaaactt aagcttgcca ccatgtccaa aggagaagaa ctgtttaccg gtgttgtgcc
      961 aattttggtt gaactcgatg gtgatgtcaa cggacataag ttctcagtga gaggcgaagg
     1021 agaaggtgac gccaccattg gaaaattgac tcttaaattc atctgtacta ctggtaaact
     1081 tcctgtacca tggccgactc tcgtaacaac gcttacgtac ggagttcagt gcttttcgag
     1141 atacccagac catatgaaaa gacatgactt ttttaagtcg gctatgcctg aaggttacgt
     1201 gcaagaaaga acaatttcgt tcaaagatga tggaaaatat aaaactagag cagttgttaa
     1261 atttgaagga gatactttgg ttaaccgcat tgaactgaaa ggaacagatt ttaaagaaga
     1321 tggtaatatt cttggacaca aactcgaata caattttaat agtcataacg tatacatcac
     1381 tgctgataag caaaagaacg gaattaaagc gaatttcaca gtacgccata atgtagaaga
     1441 tggcagtgtt caacttgccg accattacca acaaaacacc cctattggag acggtccggt
     1501 acttcttcct gataatcact acctctcaac acaaacagtc ctgagcaaag atccaaatga
     1561 aaaaggaggc agcggatcca ccactaacat tgcgccggct ctggacggtg cgtttcatcg
     1621 tatcacttct tcatcctccc tgatgggcct gtgtgacgaa accgcgacta ttatccgtga
     1681 gattactggc tacgaccgtg tgatggtagt acgtttcgat gaagagggta atgccgaaat
     1741 tctgtccgaa cgtcgtcgtg cggacctgga agcgttcctg ggtaaccgct acccggcgtc
     1801 tactattccg cagatcgctc gtcgcctgta cgaacataac cgtgttcgcc tgctggtaga
     1861 tgtgaactat actccggttc cgctacagcc gcgcatcagc ccgctgaacg gtcgtgatct
     1921 ggatatgtcc ctgtcttgcc tgcgctctat gtccccggtc caccagaaat acatgcagga
     1981 catgggcgtt ggcgcgaccc tggtttgctc tctgatggtg tctggtcgtt tgtggggtct
     2041 gatcgcttgc caccactacg aaccgcgctt cgttccgttc cacattcgcg ctgctggcga
     2101 agcgctggcg gaaacttgtg cgaaccgcat cgcgacgctg gagagcgaaa atctctactt
     2161 ccagagtgaa ttctgtgaac gcgagcagat ccacctggcg ggctccattc agccgcacgg
     2221 tatcctgctg gctgtgaaag agccagacaa cgtggtgatc caggcttcta tcaacgctgc
     2281 ggagttcctg aacaccaact ctgttgttgg ccgtccgctg cgtgacctgg gcggcgatct
     2341 gcctttgcag atcctgccgc acctgaacgg cccgctgcac ctggctccgg tgaccctgcg
     2401 ttgtaccgtg ggttctccgc cgcgccgtgt ggactgtacc attcaccgtc cgtctaacgg
     2461 cggcctgatc gtagaactgg aaccagcaac caagctcgag ggaggcagcc gggaccacat
     2521 ggtgctgcac gagtacgtga acgccgccgg catcacataa tctagagggc ccgtttaaac
     2581 ccgctgatca gcctcgactg tgccttctag ttgccagcca tctgttgttt gcccctcccc
     2641 cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc ctttcctaat aaaatgagga
     2701 aattgcatcg cattgtctga gtaggtgtca ttctattctg gggggtgggg tggggcagga
     2761 cagcaagggg gaggattggg aagacaatag caggcatgct ggggatgcgg tgggctctat
     2821 ggcttctgag gcggaaagaa ccagctgggg ctctaggggg tatccccacg cgccctgtag
     2881 cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag
     2941 cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt
     3001 tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg ctttacggca
     3061 cctcgacccc aaaaaacttg attagggtga tggttcacgt agtgggccat cgccctgata
     3121 gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac tcttgttcca
     3181 aactggaaca acactcaacc ctatctcggt ctattctttt gatttataag ggattttgcc
     3241 gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg cgaattaatt
     3301 ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag gctccccagc aggcagaagt
     3361 atgcaaagca tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc aggctcccca
     3421 gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccatagt cccgccccta
     3481 actccgccca tcccgcccct aactccgccc agttccgccc attctccgcc ccatggctga
     3541 ctaatttttt ttatttatgc agaggccgag gccgcctctg cctctgagct attccagaag
     3601 tagtgaggag gcttttttgg aggcctaggc ttttgcaaaa agctcccggg agcttgtata
     3661 tccattttcg gatctgatca agagacagga tgaggatcgt ttcgcatgat tgaacaagat
     3721 ggattgcacg caggttctcc ggccgcttgg gtggagaggc tattcggcta tgactgggca
     3781 caacagacaa tcggctgctc tgatgccgcc gtgttccggc tgtcagcgca ggggcgcccg
     3841 gttctttttg tcaagaccga cctgtccggt gccctgaatg aactgcagga cgaggcagcg
     3901 cggctatcgt ggctggccac gacgggcgtt ccttgcgcag ctgtgctcga cgttgtcact
     3961 gaagcgggaa gggactggct gctattgggc gaagtgccgg ggcaggatct cctgtcatct
     4021 caccttgctc ctgccgagaa agtatccatc atggctgatg caatgcggcg gctgcatacg
     4081 cttgatccgg ctacctgccc attcgaccac caagcgaaac atcgcatcga gcgagcacgt
     4141 actcggatgg aagccggtct tgtcgatcag gatgatctgg acgaagagca tcaggggctc
     4201 gcgccagccg aactgttcgc caggctcaag gcgcgcatgc ccgacggcga ggatctcgtc
     4261 gtgacccatg gcgatgcctg cttgccgaat atcatggtgg aaaatggccg cttttctgga
     4321 ttcatcgact gtggccggct gggtgtggcg gaccgctatc aggacatagc gttggctacc
     4381 cgtgatattg ctgaagagct tggcggcgaa tgggctgacc gcttcctcgt gctttacggt
     4441 atcgccgctc ccgattcgca gcgcatcgcc ttctatcgcc ttcttgacga gttcttctga
     4501 gcgggactct ggggttcgaa atgaccgacc aagcgacgcc caacctgcca tcacgagatt
     4561 tcgattccac cgccgccttc tatgaaaggt tgggcttcgg aatcgttttc cgggacgccg
     4621 gctggatgat cctccagcgc ggggatctca tgctggagtt cttcgcccac cccaacttgt
     4681 ttattgcagc ttataatggt tacaaataaa gcaatagcat cacaaatttc acaaataaag
     4741 catttttttc actgcattct agttgtggtt tgtccaaact catcaatgta tcttatcatg
     4801 tctgtatacc gtcgacctct agctagagct tggcgtaatc atggtcatag ctgtttcctg
     4861 tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagc ataaagtgta
     4921 aagcctgggg tgcctaatga gtgagctaac tcacattaat tgcgttgcgc tcactgcccg
     4981 ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga
     5041 gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg
     5101 tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag
     5161 aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc
     5221 gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca
     5281 aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt
     5341 ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc
     5401 tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc
     5461 tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc
     5521 ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact
     5581 tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg
     5641 ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta
     5701 tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca
     5761 aacaaaccac cgctggtagc ggtttttttg tttgcaagca gcagattacg cgcagaaaaa
     5821 aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa
     5881 actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt
     5941 taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca
     6001 gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca
     6061 tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc
     6121 ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa
     6181 accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc
     6241 agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca
     6301 acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat
     6361 tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag
     6421 cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac
     6481 tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt
     6541 ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt
     6601 gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc
     6661 tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat
     6721 ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca
     6781 gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga
     6841 cacggaaatg ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg
     6901 gttattgtct catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg
     6961 ttccgcgcac atttccccga aaagtgccac ctgacgtc
//
