LOCUS       sequence-352463-        5052 bp    DNA     circular SYN 14-APR-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5052)
  AUTHORS   Hammond GR, Fischer MJ, Anderson KE, Holdich J, Koteci A, Balla T, 
            Irvine RF
  TITLE     PI4P and PI(4,5)P2 Are Essential But Independent Lipid Determinants 
            of Membrane Identity.
  JOURNAL   Science. 2012 Jun 21.
  PUBMED    22722250
REFERENCE   2  (bases 1 to 5052)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Science. 
            2012 Jun 21."
COMMENT     Sequence Label: LYN11-FRB-mcherry
FEATURES             Location/Qualifiers
     source          1..5052
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        136..327
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     284..304
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        complement(329..433)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      460..915
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(547..566)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     757..778
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     polyA_signal    922..1043
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(933..952)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     987..1006
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(1165..1875)
                     /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(1443..1462)
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     1712..1730
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     regulatory      1872..1881
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             complement(1888..1914)
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             complement(1921..2199)
                     /codon_start=1
                     /product="FKBP-rapamycin binding domain of human FRAP"
                     /label=FRB
                     /translation="ILWHEMWHEGLEEASRLYFGERNVKGMFEVLEPLHAMMERGPQTL
                     KETSFNQAYGRDLMEAQEWCRKYMKSGNVKDLTQAWDLYYHVFRRISK"
     CDS             2214..2231
                     /codon_start=1
                     /product="HQHQHQ metal affinity tag"
                     /label=HQ tag
                     /note="The HQ tag binds to nickel and other metal ions much
                     like the 6xHis tag, but may allow elution at lower 
                     concentrations of imidazole (Godat et al., 2008)."
                     /translation="HQHQHQ"
     promoter        complement(2315..2518)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(2344..2364)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        2519..2822
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     rep_origin      complement(2997..3585)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3077..3096)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     polyA_signal    3914..3961
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     primer_bind     3986..4005
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     CDS             complement(4193..4987)
                     /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             complement(4193..4987)
                     /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(4304..4323)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     4914..4933
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(join(5022..5052,1..327))
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      join(5036..5052,1..119)
                     /label=SV40 ori
                     /note="SV40 origin of replication"
ORIGIN
        1 cctcactact tctggaatag ctcagaggcc gaggcggcct cggcctctgc ataaataaaa
       61 aaaattagtc agccatgggg cggagaatgg gcggaactgg gcggagttag gggcgggatg
      121 ggcggagtta ggggcgggac tatggttgct gactaattga gatgcatgct ttgcatactt
      181 ctgcctgctg gggagcctgg ggactttcca cacctggttg ctgactaatt gagatgcatg
      241 ctttgcatac ttctgcctgc tggggagcct ggggactttc cacaccctaa ctgacacaca
      301 ttccacagct ggttctttcc gcctcaggac tcttcctttt tcaatattat tgaagcattt
      361 atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa
      421 taggggttcc gcgcacattt ccccgaaaag tgccacctga cgcgccctgt agcggcgcat
      481 taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc agcgccctag
      541 cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc tttccccgtc
      601 aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg cacctcgacc
      661 ccaaaaaact tgattagggt gatggttcac gtagtgggcc atcgccctga tagacggttt
      721 ttcgcccttt gacgttggag tccacgttct ttaatagtgg actcttgttc caaactggaa
      781 caacactcaa ccctatctcg gtctattctt ttgatttata agggattttg ccgatttcgg
      841 cctattggtt aaaaaatgag ctgatttaac aaaaatttaa cgcgaatttt aacaaaatat
      901 taacgcttac aatttacgcc ttaagataca ttgatgagtt tggacaaacc acaactagaa
      961 tgcagtgaaa aaaatgcttt atttgtgaaa tttgtgatgc tattgcttta tttgtaacca
     1021 ttataagctg caataaacaa gttaacaaca acaattgcat tcattttatg tttcaggttc
     1081 agggggaggt gtgggaggtt ttttaaagca agtaaaacct ctacaaatgt ggtatggctg
     1141 attatgatct agagtcgcgg ccgcttactt gtacagctcg tccatgccgc cggtggagtg
     1201 gcggccctcg gcgcgttcgt actgttccac gatggtgtag tcctcgttgt gggaggtgat
     1261 gtccaacttg atgttgacgt tgtaggcgcc gggcagctgc acgggcttct tggccttgta
     1321 ggtggtcttg acctcagcgt cgtagtggcc gccgtccttc agcttcagcc tctgcttgat
     1381 ctcgcccttc agggcgccgt cctcggggta catccgctcg gaggaggcct cccagcccat
     1441 ggtcttcttc tgcattacgg ggccgtcgga ggggaagttg gtgccgcgca gcttcacctt
     1501 gtagatgaac tcgccgtcct gcagggagga gtcctgggtc acggtcacca cgccgccgtc
     1561 ctcgaagttc atcacgcgct cccacttgaa gccctcgggg aaggacagct tcaagtagtc
     1621 ggggatgtcg gcggggtgct tcacgtaggc cttggagccg tacatgaact gaggggacag
     1681 gatgtcccag gcgaagggca gggggccacc cttggtcacc ttcagcttgg cggtctgggt
     1741 gccctcgtag gggcggccct cgccctcgcc ctcgatctcg aactcgtggc cgttcacgga
     1801 gccctccatg tgcaccttga agcgcatgaa ctccttgatg atggccatgt tatcctcctc
     1861 gcccttgctc accatggtgg cgaccggtgc gtagtctggt acgtcgtacg gataactagt
     1921 ctttgagatt cgtcggaaca catgataata gaggtcccag gcttgggtga ggtccttgac
     1981 attccctgat ttcatgtact tcctgcacca ctcttgggcc tccattaaat ctcgaccata
     2041 ggcctgatta aaggatgttt ccttcagagt ctgggggccc cgttccatca tagcatgcaa
     2101 gggctccagc acctcaaaca tgcctttcac gttcctttcc ccaaagtaca aacgagatgc
     2161 ctcttccagg ccttcatgcc acatctcatg ccagaggatt ctagatagga tagcaccagc
     2221 accagcacca gcacctgaat tcgcagatct gctgtctttc ccttttgatt ttatacatcc
     2281 catggtgcta gcggatctga cggttcacta aaccagctct gcttatatag acctcccacc
     2341 gtacacgcct accgcccatt tgcgtcaatg gggcggagtt gttacgacat tttggaaagt
     2401 cccgttgatt ttggtgccaa aacaaactcc cattgacgtc aatggggtgg agacttggaa
     2461 atccccgtga gtcaaaccgc tatccacgcc cattgatgta ctgccaaaac cgcatcacca
     2521 tggtaatagc gatgactaat acgtagatgt actgccaagt aggaaagtcc cataaggtca
     2581 tgtactgggc ataatgccag gcgggccatt taccgtcatt gacgtcaata gggggcgtac
     2641 ttggcatatg atacacttga tgtactgcca agtgggcagt ttaccgtaaa tactccaccc
     2701 attgacgtca atggaaagtc cctattggcg ttactatggg aacatacgtc attattgacg
     2761 tcaatgggcg ggggtcgttg ggcggtcagc caggcgggcc atttaccgta agttatgtaa
     2821 cgcggaactc catatatggg ctatgaacta atgaccccgt aattgattac tattaataac
     2881 taatgcatgg cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg
     2941 tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc
     3001 cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga
     3061 aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct
     3121 cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg
     3181 gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag
     3241 ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat
     3301 cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac
     3361 aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac
     3421 tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc agttaccttc
     3481 ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt
     3541 tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc
     3601 ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg
     3661 agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca
     3721 atctaaagta tatatgagta acctgaggct atggcagggc ctgccgcccc gacgttggct
     3781 gcgagccctg ggccttcacc cgaacttggg gggtggggtg gggaaaagga agaaacgcgg
     3841 gcgtattggc cccaatgggg tctcggtggg gtatcgacag agtgccagcc ctgggaccga
     3901 accccgcgtt tatgaacaaa cgacccaaca ccgtgcgttt tattctgtct ttttattgcc
     3961 gtcatagcgc gggttccttc cggtattgtc tccttccgtg tttcagttag cctcccccta
     4021 gggtgggcga agaactccag catgagatcc ccgcgctgga ggatcatcca gccggcgtcc
     4081 cggaaaacga ttccgaagcc caacctttca tagaaggcgg cggtggaatc gaaatctcgt
     4141 gatggcaggt tgggcgtcgc ttggtcggtc atttcgaacc ccagagtccc gctcagaaga
     4201 actcgtcaag aaggcgatag aaggcgatgc gctgcgaatc gggagcggcg ataccgtaaa
     4261 gcacgaggaa gcggtcagcc cattcgccgc caagctcttc agcaatatca cgggtagcca
     4321 acgctatgtc ctgatagcgg tccgccacac ccagccggcc acagtcgatg aatccagaaa
     4381 agcggccatt ttccaccatg atattcggca agcaggcatc gccatgggtc acgacgagat
     4441 cctcgccgtc gggcatgctc gccttgagcc tggcgaacag ttcggctggc gcgagcccct
     4501 gatgctcttc gtccagatca tcctgatcga caagaccggc ttccatccga gtacgtgctc
     4561 gctcgatgcg atgtttcgct tggtggtcga atgggcaggt agccggatca agcgtatgca
     4621 gccgccgcat tgcatcagcc atgatggata ctttctcggc aggagcaagg tgagatgaca
     4681 ggagatcctg ccccggcact tcgcccaata gcagccagtc ccttcccgct tcagtgacaa
     4741 cgtcgagcac agctgcgcaa ggaacgcccg tcgtggccag ccacgatagc cgcgctgcct
     4801 cgtcttgcag ttcattcagg gcaccggaca ggtcggtctt gacaaaaaga accgggcgcc
     4861 cctgcgctga cagccggaac acggcggcat cagagcagcc gattgtctgt tgtgcccagt
     4921 catagccgaa tagcctctcc acccaagcgg ccggagaacc tgcgtgcaat ccatcttgtt
     4981 caatcatgcg aaacgatcct catcctgtct cttgatcgat ctttgcaaaa gcctaggcct
     5041 ccaaaaaagc ct
//
