LOCUS       sequence-60932-e        6236 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 6236)
  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 6236)
  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: pIC111
FEATURES             Location/Qualifiers
     source          1..6236
                     /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             join(1079..1081,1082..1084,1085..1798)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(1079..1081,1082..1084,1085..1795)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1124..1145)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1385..1404)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1732..1753
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(1830..1847)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1836..2060
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2106..2534
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2193..2212)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2403..2424
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2543..2563)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2548..2877
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2728..2863
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2790..2809
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2944..3738
                     /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(2998..3017)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3608..3627
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3912..4033
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3949..3968)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4003..4022
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4082..4098)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4082..4098)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4095..4117)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4106..4122
                     /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(4130..4136,4137..4154,4155..4160))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4175..4196
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4313..4330)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4484..5069)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4564..4583)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5240..6031,6032..6100))
                     /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     5863..5882
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6101..6205)
                     /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 ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg agctggacgg
     1141 cgacgtaaac ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg ccacctacgg
     1201 caagctgacc ctgaagttca tctgcaccac cggcaagctg cccgtgccct ggcccaccct
     1261 cgtgaccacc ctgacctacg gcgtgcagtg cttcagccgc taccccgacc acatgaagca
     1321 gcacgacttc ttcaagtccg ccatgcccga aggctacgtc caggagcgca ccatcttctt
     1381 caaggacgac ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg acaccctggt
     1441 gaaccgcatc gagctgaagg gcatcgactt caaggaggac ggcaacatcc tggggcacaa
     1501 gctggagtac aactacaaca gccacaacgt ctatatcatg gccgacaagc agaagaacgg
     1561 catcaaggtg aacttcaaga tccgccacaa catcgaggac ggcagcgtgc agctcgccga
     1621 ccactaccag cagaacaccc ccatcggcga cggccccgtg ctgctgcccg acaaccacta
     1681 cctgagcacc cagtccgccc tgagcaaaga ccccaacgag aagcgcgatc acatggtcct
     1741 gctggagttc gtgaccgccg ccgggatcac tctcggcatg gacgagctgt acaagtaatc
     1801 tagagggccc gtttaaaccc gctgatcagc ctcgactgtg ccttctagtt gccagccatc
     1861 tgttgtttgc ccctcccccg tgccttcctt gaccctggaa ggtgccactc ccactgtcct
     1921 ttcctaataa aatgaggaaa ttgcatcgca ttgtctgagt aggtgtcatt ctattctggg
     1981 gggtggggtg gggcaggaca gcaaggggga ggattgggaa gacaatagca ggcatgctgg
     2041 ggatgcggtg ggctctatgg cttctgaggc ggaaagaacc agctggggct ctagggggta
     2101 tccccacgcg ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt
     2161 gaccgctaca cttgccagcg ccctagcgcc cgctcctttc gctttcttcc cttcctttct
     2221 cgccacgttc gccggctttc cccgtcaagc tctaaatcgg gggctccctt tagggttccg
     2281 atttagtgct ttacggcacc tcgaccccaa aaaacttgat tagggtgatg gttcacgtag
     2341 tgggccatcg ccctgataga cggtttttcg ccctttgacg ttggagtcca cgttctttaa
     2401 tagtggactc ttgttccaaa ctggaacaac actcaaccct atctcggtct attcttttga
     2461 tttataaggg attttgccga tttcggccta ttggttaaaa aatgagctga tttaacaaaa
     2521 atttaacgcg aattaattct gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc
     2581 tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac caggtgtgga
     2641 aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca
     2701 accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag ttccgcccat
     2761 tctccgcccc atggctgact aatttttttt atttatgcag aggccgaggc cgcctctgcc
     2821 tctgagctat tccagaagta gtgaggaggc ttttttggag gcctaggctt ttgcaaaaag
     2881 ctcccgggag cttgtatatc cattttcgga tctgatcaag agacaggatg aggatcgttt
     2941 cgcatgattg aacaagatgg attgcacgca ggttctccgg ccgcttgggt ggagaggcta
     3001 ttcggctatg actgggcaca acagacaatc ggctgctctg atgccgccgt gttccggctg
     3061 tcagcgcagg ggcgcccggt tctttttgtc aagaccgacc tgtccggtgc cctgaatgaa
     3121 ctgcaggacg aggcagcgcg gctatcgtgg ctggccacga cgggcgttcc ttgcgcagct
     3181 gtgctcgacg ttgtcactga agcgggaagg gactggctgc tattgggcga agtgccgggg
     3241 caggatctcc tgtcatctca ccttgctcct gccgagaaag tatccatcat ggctgatgca
     3301 atgcggcggc tgcatacgct tgatccggct acctgcccat tcgaccacca agcgaaacat
     3361 cgcatcgagc gagcacgtac tcggatggaa gccggtcttg tcgatcagga tgatctggac
     3421 gaagagcatc aggggctcgc gccagccgaa ctgttcgcca ggctcaaggc gcgcatgccc
     3481 gacggcgagg atctcgtcgt gacccatggc gatgcctgct tgccgaatat catggtggaa
     3541 aatggccgct tttctggatt catcgactgt ggccggctgg gtgtggcgga ccgctatcag
     3601 gacatagcgt tggctacccg tgatattgct gaagagcttg gcggcgaatg ggctgaccgc
     3661 ttcctcgtgc tttacggtat cgccgctccc gattcgcagc gcatcgcctt ctatcgcctt
     3721 cttgacgagt tcttctgagc gggactctgg ggttcgaaat gaccgaccaa gcgacgccca
     3781 acctgccatc acgagatttc gattccaccg ccgccttcta tgaaaggttg ggcttcggaa
     3841 tcgttttccg ggacgccggc tggatgatcc tccagcgcgg ggatctcatg ctggagttct
     3901 tcgcccaccc caacttgttt attgcagctt ataatggtta caaataaagc aatagcatca
     3961 caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca
     4021 tcaatgtatc ttatcatgtc tgtataccgt cgacctctag ctagagcttg gcgtaatcat
     4081 ggtcatagct gtttcctgtg tgaaattgtt atccgctcac aattccacac aacatacgag
     4141 ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt gagctaactc acattaattg
     4201 cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa
     4261 tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca
     4321 ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg
     4381 taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc
     4441 agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc
     4501 cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac
     4561 tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc
     4621 tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata
     4681 gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc
     4741 acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca
     4801 acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag
     4861 cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta
     4921 gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg
     4981 gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tttttttgtt tgcaagcagc
     5041 agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg
     5101 acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga
     5161 tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg
     5221 agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct
     5281 gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg
     5341 agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc
     5401 cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa
     5461 ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc
     5521 cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt
     5581 cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc
     5641 ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt
     5701 tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc
     5761 catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt
     5821 gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata
     5881 gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga
     5941 tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag
     6001 catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa
     6061 aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt
     6121 attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga
     6181 aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct gacgtc
//
