LOCUS       sequence-60661-e        4850 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 4850)
  AUTHORS   Cheeseman IM, Desai A
  TITLE     A combined approach for the localization and tandem affinity 
            purification of protein complexes from metazoans.
  JOURNAL   Sci STKE. 2005 Jan 11;2005(266):pl1.
  PUBMED    15644491
REFERENCE   2  (bases 1 to 4850)
  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"; journalName: "Sci STKE.";
            date: "2005-01-11"; volume: "2005(266)"; pages: "pl1"
COMMENT     Sequence Label: pIC194
FEATURES             Location/Qualifiers
     source          1..4850
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             58..78
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             91..135
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     misc_feature    154..206
                     /label=MCS
                     /note="multiple cloning site"
     polyA_signal    330..451
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(367..386)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     421..440
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(458..913)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(595..616)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     807..826
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        940..1044
                     /gene="bla"
                     /label=AmpR promoter
     promoter        1046..1403
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        1046..1237
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(1069..1089)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      1254..1389
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             1438..2232
                     /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             1438..2232
                     /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(1492..1511)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2102..2121
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(2420..2439)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    2464..2511
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      2840..3428
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3329..3348
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     enhancer        3603..3906
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        3907..4110
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     4061..4081
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      4149..4158
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(4155..4850,1..12)
                     /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(4300..4318)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     4568..4587
                     /label=mCherry-F
                     /note="mCherry, forward primer"
ORIGIN
        1 gagctgtaca agtccggagg ggatcggtgg agctccaccg gtggcggccg ctctagagag
       61 aatctttatt ttcagggcgc cgccaaattc aaagaaaccg ctgctgctaa attcgaacgc
      121 cagcacatgg acagcggagg tggaggtact agtctcgagc tcaagcttcg aattctgcag
      181 tcgacggtac cgcgggcccg ggatccaccg gatctagata actgatcata atcagccata
      241 ccacatttgt agaggtttta cttgctttaa aaaacctccc acacctcccc ctgaacctga
      301 aacataaaat gaatgcaatt gttgttgtta acttgtttat tgcagcttat aatggttaca
      361 aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt
      421 gtggtttgtc caaactcatc aatgtatctt aacgcgtaaa ttgtaagcgt taatattttg
      481 ttaaaattcg cgttaaattt ttgttaaatc agctcatttt ttaaccaata ggccgaaatc
      541 ggcaaaatcc cttataaatc aaaagaatag accgagatag ggttgagtgt tgttccagtt
      601 tggaacaaga gtccactatt aaagaacgtg gactccaacg tcaaagggcg aaaaaccgtc
      661 tatcagggcg atggcccact acgtgaacca tcaccctaat caagtttttt ggggtcgagg
      721 tgccgtaaag cactaaatcg gaaccctaaa gggagccccc gatttagagc ttgacgggga
      781 aagccggcga acgtggcgag aaaggaaggg aagaaagcga aaggagcggg cgctagggcg
      841 ctggcaagtg tagcggtcac gctgcgcgta accaccacac ccgccgcgct taatgcgccg
      901 ctacagggcg cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta
      961 tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt
     1021 caataatatt gaaaaaggaa gagtcctgag gcggaaagaa ccagctgtgg aatgtgtgtc
     1081 agttagggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc
     1141 tcaattagtc agcaaccagg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc
     1201 aaagcatgca tctcaattag tcagcaacca tagtcccgcc cctaactccg cccatcccgc
     1261 ccctaactcc gcccagttcc gcccattctc cgccccatgg ctgactaatt ttttttattt
     1321 atgcagaggc cgaggccgcc tcggcctctg agctattcca gaagtagtga ggaggctttt
     1381 ttggaggcct aggcttttgc aaagatcgat caagagacag gatgaggatc gtttcgcatg
     1441 attgaacaag atggattgca cgcaggttct ccggccgctt gggtggagag gctattcggc
     1501 tatgactggg cacaacagac aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg
     1561 caggggcgcc cggttctttt tgtcaagacc gacctgtccg gtgccctgaa tgaactgcaa
     1621 gacgaggcag cgcggctatc gtggctggcc acgacgggcg ttccttgcgc agctgtgctc
     1681 gacgttgtca ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc ggggcaggat
     1741 ctcctgtcat ctcaccttgc tcctgccgag aaagtatcca tcatggctga tgcaatgcgg
     1801 cggctgcata cgcttgatcc ggctacctgc ccattcgacc accaagcgaa acatcgcatc
     1861 gagcgagcac gtactcggat ggaagccggt cttgtcgatc aggatgatct ggacgaagag
     1921 catcaggggc tcgcgccagc cgaactgttc gccaggctca aggcgagcat gcccgacggc
     1981 gaggatctcg tcgtgaccca tggcgatgcc tgcttgccga atatcatggt ggaaaatggc
     2041 cgcttttctg gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata
     2101 gcgttggcta cccgtgatat tgctgaagag cttggcggcg aatgggctga ccgcttcctc
     2161 gtgctttacg gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg ccttcttgac
     2221 gagttcttct gagcgggact ctggggttcg aaatgaccga ccaagcgacg cccaacctgc
     2281 catcacgaga tttcgattcc accgccgcct tctatgaaag gttgggcttc ggaatcgttt
     2341 tccgggacgc cggctggatg atcctccagc gcggggatct catgctggag ttcttcgccc
     2401 accctagggg gaggctaact gaaacacgga aggagacaat accggaagga acccgcgcta
     2461 tgacggcaat aaaaagacag aataaaacgc acggtgttgg gtcgtttgtt cataaacgcg
     2521 gggttcggtc ccagggctgg cactctgtcg ataccccacc gagaccccat tggggccaat
     2581 acgcccgcgt ttcttccttt tccccacccc accccccaag ttcgggtgaa ggcccagggc
     2641 tcgcagccaa cgtcggggcg gcaggccctg ccatagcctc aggttactca tatatacttt
     2701 agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc ctttttgata
     2761 atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag
     2821 aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa
     2881 caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt
     2941 ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgtcctt ctagtgtagc
     3001 cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa
     3061 tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa
     3121 gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc
     3181 ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa
     3241 gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa
     3301 caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg
     3361 ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc
     3421 tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg
     3481 ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgccatgc
     3541 attagttatt aatagtaatc aattacgggg tcattagttc atagcccata tatggagttc
     3601 cgcgttacat aacttacggt aaatggcccg cctggctgac cgcccaacga cccccgccca
     3661 ttgacgtcaa taatgacgta tgttcccata gtaacgccaa tagggacttt ccattgacgt
     3721 caatgggtgg agtatttacg gtaaactgcc cacttggcag tacatcaagt gtatcatatg
     3781 ccaagtacgc cccctattga cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag
     3841 tacatgacct tatgggactt tcctacttgg cagtacatct acgtattagt catcgctatt
     3901 accatggtga tgcggttttg gcagtacatc aatgggcgtg gatagcggtt tgactcacgg
     3961 ggatttccaa gtctccaccc cattgacgtc aatgggagtt tgttttggca ccaaaatcaa
     4021 cgggactttc caaaatgtcg taacaactcc gccccattga cgcaaatggg cggtaggcgt
     4081 gtacggtggg aggtctatat aagcagagct ggtttagtga accgtcagat ccgctagcgc
     4141 taccggtcgc caccatggtg agcaagggcg aggaggataa catggccatc atcaaggagt
     4201 tcatgcgctt caaggtgcac atggagggct ccgtgaacgg ccacgagttc gagatcgagg
     4261 gcgagggcga gggccgcccc tacgagggca cccagaccgc caagctgaag gtgaccaagg
     4321 gtggccccct gcccttcgcc tgggacatcc tgtcccctca gttcatgtac ggctccaagg
     4381 cctacgtgaa gcaccccgcc gacatccccg actacttgaa gctgtccttc cccgagggct
     4441 tcaagtggga gcgcgtgatg aacttcgagg acggcggcgt ggtgaccgtg acccaggact
     4501 cctccctgca ggacggcgag ttcatctaca aggtgaagct gcgcggcacc aacttcccct
     4561 ccgacggccc cgtaatgcag aagaagacca tgggctggga ggcctcctcc gagcggatgt
     4621 accccgagga cggcgccctg aagggcgaga tcaagcagag gctgaagctg aaggacggcg
     4681 gccactacga cgctgaggtc aagaccacct acaaggccaa gaagcccgtg cagctgcccg
     4741 gcgcctacaa cgtcaacatc aagttggaca tcacctccca caacgaggac tacaccatcg
     4801 tggaacagta cgaacgcgcc gagggccgcc actccaccgg cggcatggac
//
