LOCUS       sequence-134678-        5889 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of EGFP-TEV-S-tag DSN1 for protein localization
            and affinity purification (LAP).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5889)
  TITLE     Cheeseman Lab Plasmids
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 5889)
  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: 
            "Unpublished"
COMMENT     Sequence Label: pIC80
FEATURES             Location/Qualifiers
     source          1..5889
                     /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"
     regulatory      607..616
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(613..615,616..618,619..1329)
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(658..679)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(919..938)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1266..1287
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             1375..1395
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             1408..1452
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     polyA_signal    2677..2798
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2714..2733)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2768..2787
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2805..3260)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2942..2963)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3154..3173
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3287..3391
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3393..3750
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3393..3584
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3416..3436)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3601..3736
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3785..4579
                     /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             3785..4579
                     /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(3839..3858)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4449..4468
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4767..4786)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4811..4858
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5187..5775
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5676..5695
                     /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 gctagcgcta
      601 ccggtcgcca ccatggtgag caagggcgag gagctgttca ccggggtggt gcccatcctg
      661 gtcgagctgg acggcgacgt aaacggccac aagttcagcg tgtccggcga gggcgagggc
      721 gatgccacct acggcaagct gaccctgaag ttcatctgca ccaccggcaa gctgcccgtg
      781 ccctggccca ccctcgtgac caccttcggc tacggcctgc agtgcttcgc ccgctacccc
      841 gaccacatga agcagcacga cttcttcaag tccgccatgc ccgaaggcta cgtccaggag
      901 cgcaccatct tcttcaagga cgacggcaac tacaagaccc gcgccgaggt gaagttcgag
      961 ggcgacaccc tggtgaaccg catcgagctg aagggcatcg acttcaagga ggacggcaac
     1021 atcctggggc acaagctgga gtacaactac aacagccaca acgtctatat catggccgac
     1081 aagcagaaga acggcatcaa ggtgaacttc aagatccgcc acaacatcga ggacggcagc
     1141 gtgcagctcg ccgaccacta ccagcagaac acccccatcg gcgacggccc cgtgctgctg
     1201 cccgacaacc actacctgag ctaccagtcc gccctgagca aagaccccaa cgagaagcgc
     1261 gatcacatgg tcctgctgga gttcgtgacc gccgccggga tcactctcgg catggacgag
     1321 ctgtacaagt ccggagggga tcggtggagc tccaccggtg gcggccgctc tagagagaat
     1381 ctttattttc agggcgccgc caaattcaaa gaaaccgctg ctgctaaatt cgaacgccag
     1441 cacatggaca gcggaggtgg aggtactagt ctcgagatga cttcagtgac tagatcagag
     1501 atcatagatg aaaaaggacc agtgatgtct aagactcatg atcatcaatt ggaatcaagt
     1561 ctcagtcctg tggaagtgtt tgctaaaaca tctgcctccc tggagatgaa tcaaggcgtt
     1621 tcagaggaaa gaattcacct tggctctagc cctaaaaaag ggggaaattg tgatctcagc
     1681 caccaggaaa gacttcagtc gaagtccctt catttgtctc ctcaagaaca atctgccagt
     1741 tatcaagaca ggaggcaatc ctggcggcga gcaagtatga aagaaacgaa ccggcggaag
     1801 tcgctgcatc ccattcacca gggcatcaca gagctcagcc ggtctatcag tgtcgattta
     1861 gcagaaagca aacggcttgg ctgtctcctg ctttccagtt tccagttctc tattcagaaa
     1921 cttgaacctt tcctaaggga cactaagggc ttcagtcttg aaagttttag agccaaagca
     1981 tcttctcttt ctgaagaatt gaaacatttt gcagacggac tggaaactga tggaactcta
     2041 caaaaatgtt ttgaagattc aaatggaaaa gcatcagatt tttctttgga agcatctgtg
     2101 gctgagatga aggaatacat aacaaagttt tctttagaac gtcagacttg ggatcagctc
     2161 ttgcttcact accagcagga ggctaaagag atattgtcca gaggatcaac tgaggccaaa
     2221 attactgagg tcaaagtgga acctatgaca tatcttgggt cttctcagaa tgaagttctt
     2281 aatacaaaac ctgactacca gaaaatatta cagaaccaga gcaaagtctt tgactgtatg
     2341 gagttggtga tggatgaact gcaaggatca gtgaaacagc tgcaggcctt tatggatgaa
     2401 agtacccagt gcttccagaa ggtgtcagta cagctcggaa agagaagcat gcaacaatta
     2461 gatccctcac cagctcgaaa actgttgaag cttcagctac agaacccacc tgccatacat
     2521 ggatctggat ctggatcttg tcagtgagga tccaccggat ctagataact gatcataatc
     2581 agccatacca catttgtaga ggttttactt gctttaaaaa acctcccaca cctccccctg
     2641 aacctgaaac ataaaatgaa tgcaattgtt gttgttaact tgtttattgc agcttataat
     2701 ggttacaaat aaagcaatag catcacaaat ttcacaaata aagcattttt ttcactgcat
     2761 tctagttgtg gtttgtccaa actcatcaat gtatcttaag gcgtaaattg taagcgttaa
     2821 tattttgtta aaattcgcgt taaatttttg ttaaatcagc tcatttttta accaataggc
     2881 cgaaatcggc aaaatccctt ataaatcaaa agaatagacc gagatagggt tgagtgttgt
     2941 tccagtttgg aacaagagtc cactattaaa gaacgtggac tccaacgtca aagggcgaaa
     3001 aaccgtctat cagggcgatg gcccactacg tgaaccatca ccctaatcaa gttttttggg
     3061 gtcgaggtgc cgtaaagcac taaatcggaa ccctaaaggg agcccccgat ttagagcttg
     3121 acggggaaag ccggcgaacg tggcgagaaa ggaagggaag aaagcgaaag gagcgggcgc
     3181 tagggcgctg gcaagtgtag cggtcacgct gcgcgtaacc accacacccg ccgcgcttaa
     3241 tgcgccgcta cagggcgcgt caggtggcac ttttcgggga aatgtgcgcg gaacccctat
     3301 ttgtttattt ttctaaatac attcaaatat gtatccgctc atgagacaat aaccctgata
     3361 aatgcttcaa taatattgaa aaaggaagag tcctgaggcg gaaagaacca gctgtggaat
     3421 gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc
     3481 atgcatctca attagtcagc aaccaggtgt ggaaagtccc caggctcccc agcaggcaga
     3541 agtatgcaaa gcatgcatct caattagtca gcaaccatag tcccgcccct aactccgccc
     3601 atcccgcccc taactccgcc cagttccgcc cattctccgc cccatggctg actaattttt
     3661 tttatttatg cagaggccga ggccgcctcg gcctctgagc tattccagaa gtagtgagga
     3721 ggcttttttg gaggcctagg cttttgcaaa gatcgatcaa gagacaggat gaggatcgtt
     3781 tcgcatgatt gaacaagatg gattgcacgc aggttctccg gccgcttggg tggagaggct
     3841 attcggctat gactgggcac aacagacaat cggctgctct gatgccgccg tgttccggct
     3901 gtcagcgcag gggcgcccgg ttctttttgt caagaccgac ctgtccggtg ccctgaatga
     3961 actgcaagac gaggcagcgc ggctatcgtg gctggccacg acgggcgttc cttgcgcagc
     4021 tgtgctcgac gttgtcactg aagcgggaag ggactggctg ctattgggcg aagtgccggg
     4081 gcaggatctc ctgtcatctc accttgctcc tgccgagaaa gtatccatca tggctgatgc
     4141 aatgcggcgg ctgcatacgc ttgatccggc tacctgccca ttcgaccacc aagcgaaaca
     4201 tcgcatcgag cgagcacgta ctcggatgga agccggtctt gtcgatcagg atgatctgga
     4261 cgaagagcat caggggctcg cgccagccga actgttcgcc aggctcaagg cgagcatgcc
     4321 cgacggcgag gatctcgtcg tgacccatgg cgatgcctgc ttgccgaata tcatggtgga
     4381 aaatggccgc ttttctggat tcatcgactg tggccggctg ggtgtggcgg accgctatca
     4441 ggacatagcg ttggctaccc gtgatattgc tgaagagctt ggcggcgaat gggctgaccg
     4501 cttcctcgtg ctttacggta tcgccgctcc cgattcgcag cgcatcgcct tctatcgcct
     4561 tcttgacgag ttcttctgag cgggactctg gggttcgaaa tgaccgacca agcgacgccc
     4621 aacctgccat cacgagattt cgattccacc gccgccttct atgaaaggtt gggcttcgga
     4681 atcgttttcc gggacgccgg ctggatgatc ctccagcgcg gggatctcat gctggagttc
     4741 ttcgcccacc ctagggggag gctaactgaa acacggaagg agacaatacc ggaaggaacc
     4801 cgcgctatga cggcaataaa aagacagaat aaaacgcacg gtgttgggtc gtttgttcat
     4861 aaacgcgggg ttcggtccca gggctggcac tctgtcgata ccccaccgag accccattgg
     4921 ggccaatacg cccgcgtttc ttccttttcc ccaccccacc ccccaagttc gggtgaaggc
     4981 ccagggctcg cagccaacgt cggggcggca ggccctgcca tagcctcagg ttactcatat
     5041 atactttaga ttgatttaaa acttcatttt taatttaaaa ggatctaggt gaagatcctt
     5101 tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac
     5161 cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc
     5221 ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca
     5281 actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgtccttcta
     5341 gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct
     5401 ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg
     5461 gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc
     5521 acacagccca gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta
     5581 tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg
     5641 gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt
     5701 cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg
     5761 cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg
     5821 ccttttgctc acatgttctt tcctgcgtta tcccctgatt ctgtggataa ccgtattacc
     5881 gccatgcat
//
