LOCUS       Exported                6892 bp ds-DNA     circular SYN 25-NOV-2020
DEFINITION  .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6892)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Wednesday, Nov 25, 2020 from SnapGene 4.3.6
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6892
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
                     /note="color: #ffffff"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
                     /note="color: #ffffff; direction: RIGHT"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
                     /note="color: #ffffff; direction: RIGHT"
     misc_feature    901..906
                     /label=Kozak
                     /note="color: #a6acb3"
     misc_feature    907..963
                     /label=MPKK-NES
                     /note="color: #ff9900; direction: RIGHT"
     CDS             979..1704
                     /codon_start=1
                     /product="photocleavable protein that separates into two 
                     fragments upon exposure to violet light (Zhang et al., 
                     2017)"
                     /label=PhoCl2f
                     /note="color: #993366"
                     /translation="VIPDYFKQSFPEGYSWERSMTYEDGGICIATNDITMEGDSFINKI
                     HFQGTNFPPNGPVMQKRTVGWEASTEKMYERDGVLKGDVKMKLLLKGGGHYRCDYRTTY
                     KVKQKPVKLPDCHFVDHRIEILSHDKDYNKVKLYEHAVAKTSTDSMDELYKGGSGGMVS
                     KGEETITSVIKPDMKNKLRMEGNVNGHAFVIEGEGSGKPFEGSQTIDLEVKEGAPLPFA
                     YDILTTAFHYGNRVFTKYPR"
     primer_bind     979..993
                     /label=Gi-pB-fw-XhoI-G-PhoCL
                     /note="color: black; sequence: 
                     GTACGACGATGACGATAAGGATCCGAGCTCGAGGATGGTGATCCCTGACTAC; 
                     added: 2019-06-20"
     primer_bind     complement(1684..1733)
                     /label=Gi-RV-PhoCl*-KpnI-pB
                     /note="color: black; sequence: 
                     CATCCGCCAAAACAGCCAAGCTTGGTACCTTAccgtgggtacttggtgaacac; 
                     added: 2019-06-20"
     CDS             1741..2448
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /note="color: #ff0000"
                     /translation="VSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEGT
                     QTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFE
                     DGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALKG
                     EIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERAE
                     GRHSTGGMDELYK"
     polyA_signal    2492..2716
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
                     /note="color: #a6acb3"
     rep_origin      2762..3190
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
                     /note="color: #ffff00"
     promoter        3204..3533
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
                     /note="color: #ffffff; direction: RIGHT"
     rep_origin      3384..3519
                     /label=SV40 ori
                     /note="SV40 origin of replication"
                     /note="color: #ffff00"
     CDS             3600..4394
                     /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))"
                     /note="color: #ccffcc"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     polyA_signal    4568..4689
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
                     /note="color: #a6acb3"
     primer_bind     complement(4738..4754)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
                     /note="color: #a020f0; direction: LEFT"
     protein_bind    4762..4778
                     /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)."
                     /note="color: #31849b"
     promoter        complement(4786..4816)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
                     /note="This reverse directional feature has 3 segments:
                        1: 4786 .. 4792 / #ffffff / -10
                        2: 4793 .. 4810 / #ffffff
                        3: 4811 .. 4816 / #ffffff / -35"
     protein_bind    4831..4852
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
                     /note="color: #31849b"
     rep_origin      complement(5140..5725)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
                     /note="color: #ffff00"
     CDS             complement(5896..6756)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /note="This feature has 2 segments:
                        1: 5896 .. 6687 / #ccffcc
                        2: 6688 .. 6756 / #ccffcc / signal sequence
                     Cleavage site after base 6687"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(6757..6861)
                     /gene="bla"
                     /label=AmpR promoter
                     /note="color: #ffffff; direction: LEFT"
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 gccaccatga acctggtgga cctgcagaag aagctggagg agctggagct ggacgagcag
      961 cagggaggca agcttagcgt gatccctgac tacttcaagc agagcttccc cgagggctac
     1021 agctgggagc gcagcatgac ctacgaggac ggcggcatct gcatcgccac caacgacatc
     1081 acaatggagg gggacagctt catcaacaag atccacttcc agggcacgaa cttccccccc
     1141 aacggccccg tgatgcagaa gaggaccgtg ggctgggagg ccagcaccga gaagatgtac
     1201 gagcgcgacg gcgtgctgaa gggcgacgtg aagatgaagc tgctgctgaa gggcggcggc
     1261 cactatcgct gcgactaccg caccacctac aaggtcaagc agaagcccgt aaagctgccc
     1321 gactgccact tcgtggacca ccgcatcgag atcctgagcc acgacaagga ctacaacaag
     1381 gtgaagctgt acgagcacgc cgtggccaag acttccaccg acagcatgga cgagctgtac
     1441 aagggtggca gcggtggcat ggtgagcaag ggcgaggaga ccattacaag cgtgatcaag
     1501 cctgacatga agaacaagct gcgcatggag ggcaacgtga acggccacgc cttcgtgatc
     1561 gagggcgagg gcagcggcaa gcccttcgag ggctctcaga cgattgattt ggaggtgaag
     1621 gagggcgccc cgctgccctt cgcctacgac atcctgacca ccgccttcca ctacggcaac
     1681 cgcgtgttca ccaagtaccc acggggtacc ggaggcgggg gcagcggagg cgggggcagc
     1741 gtgagcaagg gcgaggagga taacatggcc atcatcaagg agttcatgcg cttcaaggtg
     1801 cacatggagg gctccgtgaa cggccacgag ttcgagatcg agggcgaggg cgagggccgc
     1861 ccctacgagg gcacccagac cgccaagctg aaggtgacca agggtggccc cctgcccttc
     1921 gcctgggaca tcctgtcccc tcagttcatg tacggctcca aggcctacgt gaagcacccc
     1981 gccgacatcc ccgactactt gaagctgtcc ttccccgagg gcttcaagtg ggagcgcgtg
     2041 atgaacttcg aggacggcgg cgtggtgacc gtgacccagg actcctccct gcaggacggc
     2101 gagttcatct acaaggtgaa gctgcgcggc accaacttcc cctccgacgg ccccgtaatg
     2161 cagaagaaga ccatgggctg ggaggcctcc tccgagcgga tgtaccccga ggacggcgcc
     2221 ctgaagggcg agatcaagca gaggctgaag ctgaaggacg gcggccacta cgacgctgag
     2281 gtcaagacca cctacaaggc caagaagccc gtgcagctgc ccggcgccta caacgtcaac
     2341 atcaagttgg acatcacctc ccacaacgag gactacacca tcgtggaaca gtacgaacgc
     2401 gccgagggcc gccactccac cggcggcatg gacgagctgt acaagtaact cgagtctaga
     2461 gggcccgttt aaacccgctg atcagcctcg actgtgcctt ctagttgcca gccatctgtt
     2521 gtttgcccct cccccgtgcc ttccttgacc ctggaaggtg ccactcccac tgtcctttcc
     2581 taataaaatg aggaaattgc atcgcattgt ctgagtaggt gtcattctat tctggggggt
     2641 ggggtggggc aggacagcaa gggggaggat tgggaagaca atagcaggca tgctggggat
     2701 gcggtgggct ctatggcttc tgaggcggaa agaaccagct ggggctctag ggggtatccc
     2761 cacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc
     2821 gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc ctttctcgcc
     2881 acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt
     2941 agtgctttac ggcacctcga ccccaaaaaa cttgattagg gtgatggttc acgtagtggg
     3001 ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt ctttaatagt
     3061 ggactcttgt tccaaactgg aacaacactc aaccctatct cggtctattc ttttgattta
     3121 taagggattt tgccgatttc ggcctattgg ttaaaaaatg agctgattta acaaaaattt
     3181 aacgcgaatt aattctgtgg aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc
     3241 cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccagg tgtggaaagt
     3301 ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca
     3361 tagtcccgcc cctaactccg cccatcccgc ccctaactcc gcccagttcc gcccattctc
     3421 cgccccatgg ctgactaatt ttttttattt atgcagaggc cgaggccgcc tctgcctctg
     3481 agctattcca gaagtagtga ggaggctttt ttggaggcct aggcttttgc aaaaagctcc
     3541 cgggagcttg tatatccatt ttcggatctg atcaagagac aggatgagga tcgtttcgca
     3601 tgattgaaca agatggattg cacgcaggtt ctccggccgc ttgggtggag aggctattcg
     3661 gctatgactg ggcacaacag acaatcggct gctctgatgc cgccgtgttc cggctgtcag
     3721 cgcaggggcg cccggttctt tttgtcaaga ccgacctgtc cggtgccctg aatgaactgc
     3781 aggacgaggc agcgcggcta tcgtggctgg ccacgacggg cgttccttgc gcagctgtgc
     3841 tcgacgttgt cactgaagcg ggaagggact ggctgctatt gggcgaagtg ccggggcagg
     3901 atctcctgtc atctcacctt gctcctgccg agaaagtatc catcatggct gatgcaatgc
     3961 ggcggctgca tacgcttgat ccggctacct gcccattcga ccaccaagcg aaacatcgca
     4021 tcgagcgagc acgtactcgg atggaagccg gtcttgtcga tcaggatgat ctggacgaag
     4081 agcatcaggg gctcgcgcca gccgaactgt tcgccaggct caaggcgcgc atgcccgacg
     4141 gcgaggatct cgtcgtgacc catggcgatg cctgcttgcc gaatatcatg gtggaaaatg
     4201 gccgcttttc tggattcatc gactgtggcc ggctgggtgt ggcggaccgc tatcaggaca
     4261 tagcgttggc tacccgtgat attgctgaag agcttggcgg cgaatgggct gaccgcttcc
     4321 tcgtgcttta cggtatcgcc gctcccgatt cgcagcgcat cgccttctat cgccttcttg
     4381 acgagttctt ctgagcggga ctctggggtt cgaaatgacc gaccaagcga cgcccaacct
     4441 gccatcacga gatttcgatt ccaccgccgc cttctatgaa aggttgggct tcggaatcgt
     4501 tttccgggac gccggctgga tgatcctcca gcgcggggat ctcatgctgg agttcttcgc
     4561 ccaccccaac ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa
     4621 tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa
     4681 tgtatcttat catgtctgta taccgtcgac ctctagctag agcttggcgt aatcatggtc
     4741 atagctgttt cctgtgtgaa attgttatcc gctcacaatt ccacacaaca tacgagccgg
     4801 aagcataaag tgtaaagcct ggggtgccta atgagtgagc taactcacat taattgcgtt
     4861 gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg
     4921 ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga
     4981 ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat
     5041 acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca
     5101 aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc
     5161 tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata
     5221 aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc
     5281 gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc
     5341 acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga
     5401 accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc
     5461 ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag
     5521 gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag
     5581 aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag
     5641 ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtttt tttgtttgca agcagcagat
     5701 tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc
     5761 tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt
     5821 cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta
     5881 aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct
     5941 atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg
     6001 cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga
     6061 tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt
     6121 atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt
     6181 taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt
     6241 tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat
     6301 gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc
     6361 cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc
     6421 cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat
     6481 gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag
     6541 aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt
     6601 accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc
     6661 ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa
     6721 gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg
     6781 aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa
     6841 taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg tc
//
