LOCUS       sequence-12148-e        5021 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5021)
  AUTHORS   Karginov AV, Ding F, Kota P, Dokholyan NV, Hahn KM
  TITLE     Engineered allosteric activation of kinases in living cells.
  JOURNAL   Nat Biotechnol. 2010 Jul;28(7):743-7. doi: 10.1038/nbt.1639. Epub 
            2010 Jun 27.
  PUBMED    20581846
REFERENCE   2  (bases 1 to 5021)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: "10.1038/nbt.1639"; 
            journalName: "Nat Biotechnol"; date: "2010-07"; volume: "28"; issue:
            "7"; pages: "743-7"
COMMENT     Sequence Label: pCherry-FRB
FEATURES             Location/Qualifiers
     source          1..5021
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     519..539
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             600..1307
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(745..763)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1013..1032
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             1356..1634
                     /codon_start=1
                     /product="T2098L mutant of FKBP-rapamycin binding domain of
                     human FRAP"
                     /label=FRB*
                     /note="binds synthetic ligands such as AP21967"
                     /translation="ILWHEMWHEGLEEASRLYFGERNVKGMFEVLEPLHAMMERGPQTL
                     KETSFNQAYGRDLMEAQEWCRKYMKSGNVKDLLQAWDLYYHVFRRISK"
     polyA_signal    1809..1930
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1846..1865)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1900..1919
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(1937..2392)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2074..2095)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2286..2305
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2419..2523
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2525..2882
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2525..2716
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2548..2568)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2733..2868
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             2917..3711
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             2917..3711
                     /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
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(2971..2990)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3581..3600
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(3899..3918)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    3943..3990
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4319..4907
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4808..4827
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcacca
      601 tggtgagcaa gggcgaggag gataacatgg ccatcatcaa ggagttcatg cgcttcaagg
      661 tgcacatgga gggctccgtg aacggccacg agttcgagat cgagggcgag ggcgagggcc
      721 gcccctacga gggcacccag accgccaagc tgaaggtgac caagggtggc cccctgccct
      781 tcgcctggga catcctgtcc cctcagttca tgtacggctc caaggcctac gtgaagcacc
      841 ccgccgacat ccccgactac ttgaagctgt ccttccccga gggcttcaag tgggagcgcg
      901 tgatgaactt cgaggacggc ggcgtggtga ccgtgaccca ggactcctcc ctgcaggacg
      961 gcgagttcat ctacaaggtg aagctgcgcg gcaccaactt cccctccgac ggccccgtaa
     1021 tgcagaagaa gaccatgggc tgggaggcct cctccgagcg gatgtacccc gaggacggcg
     1081 ccctgaaggg cgagatcaag cagaggctga agctgaagga cggcggccac tacgacgctg
     1141 aggtcaagac cacctacaag gccaagaagc ccgtgcagct gcccggcgcc tacaacgtca
     1201 acatcaagtt ggacatcacc tcccacaacg aggactacac catcgtggaa cagtacgaac
     1261 gcgccgaggg ccgccactcc accggcggca tggacgagct gtacaagtcc ggactcagat
     1321 ctcgagctca agcttcgaat tcaatggctt ctagaatcct ctggcatgag atgtggcatg
     1381 aaggcctgga agaggcatct cgtttgtact ttggggaaag gaacgtgaaa ggcatgtttg
     1441 aggtgctgga gcccttgcat gctatgatgg aacggggccc ccagactctg aaggaaacat
     1501 cctttaatca ggcctatggt cgagatttaa tggaggccca agagtggtgc aggaagtaca
     1561 tgaaatcagg gaatgtcaag gacctcctcc aagcctggga cctctattat catgtgttcc
     1621 gacgaatctc aaagactagt taaccgtacg acgtaccaga ctacgcataa gaaaagtgag
     1681 gatccaccgg atctagataa ctgatcataa tcagccatac cacatttgta gaggttttac
     1741 ttgctttaaa aaacctccca cacctccccc tgaacctgaa acataaaatg aatgcaattg
     1801 ttgttgttaa cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa
     1861 atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca
     1921 atgtatctta aggcgtaaat tgtaagcgtt aatattttgt taaaattcgc gttaaatttt
     1981 tgttaaatca gctcattttt taaccaatag gccgaaatcg gcaaaatccc ttataaatca
     2041 aaagaataga ccgagatagg gttgagtgtt gttccagttt ggaacaagag tccactatta
     2101 aagaacgtgg actccaacgt caaagggcga aaaaccgtct atcagggcga tggcccacta
     2161 cgtgaaccat caccctaatc aagttttttg gggtcgaggt gccgtaaagc actaaatcgg
     2221 aaccctaaag ggagcccccg atttagagct tgacggggaa agccggcgaa cgtggcgaga
     2281 aaggaaggga agaaagcgaa aggagcgggc gctagggcgc tggcaagtgt agcggtcacg
     2341 ctgcgcgtaa ccaccacacc cgccgcgctt aatgcgccgc tacagggcgc gtcaggtggc
     2401 acttttcggg gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat
     2461 atgtatccgc tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag
     2521 agtcctgagg cggaaagaac cagctgtgga atgtgtgtca gttagggtgt ggaaagtccc
     2581 caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccaggt
     2641 gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt
     2701 cagcaaccat agtcccgccc ctaactccgc ccatcccgcc cctaactccg cccagttccg
     2761 cccattctcc gccccatggc tgactaattt tttttattta tgcagaggcc gaggccgcct
     2821 cggcctctga gctattccag aagtagtgag gaggcttttt tggaggccta ggcttttgca
     2881 aagatcgatc aagagacagg atgaggatcg tttcgcatga ttgaacaaga tggattgcac
     2941 gcaggttctc cggccgcttg ggtggagagg ctattcggct atgactgggc acaacagaca
     3001 atcggctgct ctgatgccgc cgtgttccgg ctgtcagcgc aggggcgccc ggttcttttt
     3061 gtcaagaccg acctgtccgg tgccctgaat gaactgcaag acgaggcagc gcggctatcg
     3121 tggctggcca cgacgggcgt tccttgcgca gctgtgctcg acgttgtcac tgaagcggga
     3181 agggactggc tgctattggg cgaagtgccg gggcaggatc tcctgtcatc tcaccttgct
     3241 cctgccgaga aagtatccat catggctgat gcaatgcggc ggctgcatac gcttgatccg
     3301 gctacctgcc cattcgacca ccaagcgaaa catcgcatcg agcgagcacg tactcggatg
     3361 gaagccggtc ttgtcgatca ggatgatctg gacgaagagc atcaggggct cgcgccagcc
     3421 gaactgttcg ccaggctcaa ggcgagcatg cccgacggcg aggatctcgt cgtgacccat
     3481 ggcgatgcct gcttgccgaa tatcatggtg gaaaatggcc gcttttctgg attcatcgac
     3541 tgtggccggc tgggtgtggc ggaccgctat caggacatag cgttggctac ccgtgatatt
     3601 gctgaagagc ttggcggcga atgggctgac cgcttcctcg tgctttacgg tatcgccgct
     3661 cccgattcgc agcgcatcgc cttctatcgc cttcttgacg agttcttctg agcgggactc
     3721 tggggttcga aatgaccgac caagcgacgc ccaacctgcc atcacgagat ttcgattcca
     3781 ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga
     3841 tcctccagcg cggggatctc atgctggagt tcttcgccca ccctaggggg aggctaactg
     3901 aaacacggaa ggagacaata ccggaaggaa cccgcgctat gacggcaata aaaagacaga
     3961 ataaaacgca cggtgttggg tcgtttgttc ataaacgcgg ggttcggtcc cagggctggc
     4021 actctgtcga taccccaccg agaccccatt ggggccaata cgcccgcgtt tcttcctttt
     4081 ccccacccca ccccccaagt tcgggtgaag gcccagggct cgcagccaac gtcggggcgg
     4141 caggccctgc catagcctca ggttactcat atatacttta gattgattta aaacttcatt
     4201 tttaatttaa aaggatctag gtgaagatcc tttttgataa tctcatgacc aaaatccctt
     4261 aacgtgagtt ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt
     4321 gagatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag
     4381 cggtggtttg tttgccggat caagagctac caactctttt tccgaaggta actggcttca
     4441 gcagagcgca gataccaaat actgtccttc tagtgtagcc gtagttaggc caccacttca
     4501 agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg
     4561 ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg
     4621 cgcagcggtc gggctgaacg gggggttcgt gcacacagcc cagcttggag cgaacgacct
     4681 acaccgaact gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga
     4741 gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc
     4801 ttccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg
     4861 agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg
     4921 cggccttttt acggttcctg gccttttgct ggccttttgc tcacatgttc tttcctgcgt
     4981 tatcccctga ttctgtggat aaccgtatta ccgccatgca t
//
