LOCUS       sequence-60933-e        6227 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 6227)
  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 6227)
  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: pIC112
FEATURES             Location/Qualifiers
     source          1..6227
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     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             995..1012
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1022..1045
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             1046..1069
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             1079..1789
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEENNMAVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKMRLKLKDGGHYDAEVKTTYMAKKPVQLPGAYKTDIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(1224..1242)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1492..1511
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     1686..1709
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     primer_bind     complement(1821..1838)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1827..2051
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2097..2525
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2184..2203)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2394..2415
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2534..2554)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2539..2868
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2719..2854
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2781..2800
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2935..3729
                     /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(2989..3008)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3599..3618
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3903..4024
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3940..3959)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3994..4013
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4073..4089)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4073..4089)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4086..4108)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4097..4113
                     /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(join(4121..4127,4128..4145,4146..4151))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4166..4187
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4304..4321)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4475..5060)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4555..4574)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5231..6022,6023..6091))
                     /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     5854..5873
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6092..6196)
                     /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 aagcttggta ccgagctcgg atccactagt ccagtgtggt ggaattctgc
      961 agatatccag cacagtggcg gccgcggcag cagccatcat catcatcatc acagcagcgg
     1021 cctggaagtt ctgttccagg ggcccctgga agttctgttc caggggccct ccggagggat
     1081 ggtgagcaag ggcgaggaga ataacatggc cgtcatcaag gagttcatgc gcttcaaggt
     1141 gcgcatggag ggctccgtga acggccacga gttcgagatc gagggcgagg gcgagggccg
     1201 cccctacgag ggcacccaga ccgccaagct gaaggtgacc aagggtggcc ccctgccctt
     1261 cgcctgggac atcctgtccc ctcagttcat gtacggctcc aaggcctacg tgaagcaccc
     1321 cgccgacatc cccgactact tgaagctgtc cttccccgag ggcttcaagt gggagcgcgt
     1381 gatgaacttc gaggacggcg gcgtggtgac cgtgacccag gactcctccc tgcaggacgg
     1441 cgagttcatc tacaaggtga agctgcgcgg caccaacttc ccctccgacg gccccgtaat
     1501 gcagaagaag accatgggct gggaggcctc ctccgagcgg atgtaccccg aggacggcgc
     1561 cctgaagggc gagatcaaga tgaggctgaa gctgaaggac ggcggccact acgacgccga
     1621 ggtcaagacc acctacatgg ccaagaagcc cgtgcagctg cccggcgcct acaagaccga
     1681 catcaagctg gacatcacct cccacaacga ggactacacc atcgtggaac agtacgagcg
     1741 cgccgagggc cgccactcca ccggcggcat ggacgagctg tacaagtaat ctagagggcc
     1801 cgtttaaacc cgctgatcag cctcgactgt gccttctagt tgccagccat ctgttgtttg
     1861 cccctccccc gtgccttcct tgaccctgga aggtgccact cccactgtcc tttcctaata
     1921 aaatgaggaa attgcatcgc attgtctgag taggtgtcat tctattctgg ggggtggggt
     1981 ggggcaggac agcaaggggg aggattggga agacaatagc aggcatgctg gggatgcggt
     2041 gggctctatg gcttctgagg cggaaagaac cagctggggc tctagggggt atccccacgc
     2101 gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac
     2161 acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt
     2221 cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc
     2281 tttacggcac ctcgacccca aaaaacttga ttagggtgat ggttcacgta gtgggccatc
     2341 gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact
     2401 cttgttccaa actggaacaa cactcaaccc tatctcggtc tattcttttg atttataagg
     2461 gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc
     2521 gaattaattc tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg ctccccagca
     2581 ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca
     2641 ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc aaccatagtc
     2701 ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca ttctccgccc
     2761 catggctgac taattttttt tatttatgca gaggccgagg ccgcctctgc ctctgagcta
     2821 ttccagaagt agtgaggagg cttttttgga ggcctaggct tttgcaaaaa gctcccggga
     2881 gcttgtatat ccattttcgg atctgatcaa gagacaggat gaggatcgtt tcgcatgatt
     2941 gaacaagatg gattgcacgc aggttctccg gccgcttggg tggagaggct attcggctat
     3001 gactgggcac aacagacaat cggctgctct gatgccgccg tgttccggct gtcagcgcag
     3061 gggcgcccgg ttctttttgt caagaccgac ctgtccggtg ccctgaatga actgcaggac
     3121 gaggcagcgc ggctatcgtg gctggccacg acgggcgttc cttgcgcagc tgtgctcgac
     3181 gttgtcactg aagcgggaag ggactggctg ctattgggcg aagtgccggg gcaggatctc
     3241 ctgtcatctc accttgctcc tgccgagaaa gtatccatca tggctgatgc aatgcggcgg
     3301 ctgcatacgc ttgatccggc tacctgccca ttcgaccacc aagcgaaaca tcgcatcgag
     3361 cgagcacgta ctcggatgga agccggtctt gtcgatcagg atgatctgga cgaagagcat
     3421 caggggctcg cgccagccga actgttcgcc aggctcaagg cgcgcatgcc cgacggcgag
     3481 gatctcgtcg tgacccatgg cgatgcctgc ttgccgaata tcatggtgga aaatggccgc
     3541 ttttctggat tcatcgactg tggccggctg ggtgtggcgg accgctatca ggacatagcg
     3601 ttggctaccc gtgatattgc tgaagagctt ggcggcgaat gggctgaccg cttcctcgtg
     3661 ctttacggta tcgccgctcc cgattcgcag cgcatcgcct tctatcgcct tcttgacgag
     3721 ttcttctgag cgggactctg gggttcgaaa tgaccgacca agcgacgccc aacctgccat
     3781 cacgagattt cgattccacc gccgccttct atgaaaggtt gggcttcgga atcgttttcc
     3841 gggacgccgg ctggatgatc ctccagcgcg gggatctcat gctggagttc ttcgcccacc
     3901 ccaacttgtt tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca
     3961 caaataaagc atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat
     4021 cttatcatgt ctgtataccg tcgacctcta gctagagctt ggcgtaatca tggtcatagc
     4081 tgtttcctgt gtgaaattgt tatccgctca caattccaca caacatacga gccggaagca
     4141 taaagtgtaa agcctggggt gcctaatgag tgagctaact cacattaatt gcgttgcgct
     4201 cactgcccgc tttccagtcg ggaaacctgt cgtgccagct gcattaatga atcggccaac
     4261 gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc actgactcgc
     4321 tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt
     4381 tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg
     4441 ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg
     4501 agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat
     4561 accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta
     4621 ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct
     4681 gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc
     4741 ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa
     4801 gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg
     4861 taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact agaagaacag
     4921 tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt
     4981 gatccggcaa acaaaccacc gctggtagcg gtttttttgt ttgcaagcag cagattacgc
     5041 gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt
     5101 ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct
     5161 agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt
     5221 ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc
     5281 gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac
     5341 catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat
     5401 cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg
     5461 cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata
     5521 gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta
     5581 tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt
     5641 gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag
     5701 tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa
     5761 gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc
     5821 gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt
     5881 taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc
     5941 tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta
     6001 ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa
     6061 taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca
     6121 tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac
     6181 aaataggggt tccgcgcaca tttccccgaa aagtgccacc tgacgtc
//
