LOCUS       sequence-438039-        7440 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  FAP insert for N-terminally tagging genes of interest (original FAP 
            + 2xMYC tag).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7440)
  AUTHORS   Oppenheimer KG, Hager NA, McAtee CK, Filiztekin E, Shang C, Warnick 
            JA, Bruchez MP, Brodsky JL, Prosser DC, Kwiatkowski AV, O'Donnell AF
  TITLE     Optimization of the fluorogen-activating protein tag for 
            quantitative protein trafficking and colocalization studies in S. 
            cerevisiae.
  JOURNAL   Mol Biol Cell. 2024 Jul 1;35(7):mr5. doi: 10.1091/mbc.E24-04-0174. 
            Epub 2024 May 29.
  PUBMED    38809589
REFERENCE   2  (bases 1 to 7440)
  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: "Mol Biol 
            Cell."; date: "2024-07-1"; volume: "35(7):mr5. doi"; pages: " 
            10.1091/mbc.E24-04-0174. Epub 2024 May 29"
COMMENT     Sequence Label: pRS415-TEF1pr-NFAPorig
FEATURES             Location/Qualifiers
     source          1..7440
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(167..186)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     286..308
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(346..364)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     misc_feature    406..909
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     promoter        946..1050
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1051..1119,1120..1911)
                     /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     complement(1269..1288)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2082..2670
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2571..2590
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2824..2841
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    2958..2979
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(2994..2999,3000..3017,3018..3024)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3032..3048
                     /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)."
     primer_bind     3037..3059
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     3056..3072
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     3056..3072
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     3091..3111
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        3093..3111
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        3129..3527
                     /gene="S. cerevisiae TEF1"
                     /label=TEF1 promoter
                     /note="promoter for EF-1-alpha"
     primer_bind     3531..3547
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     3531..3547
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     CDS             3552..4271
                     /codon_start=1
                     /product="fluorogen-activating protein composed of a dimer 
                     of the L5-MG single-chain antibody fragment (Szent-Gyorgyi 
                     et al., 2008)"
                     /label=FAP-alpha-2
                     /note="binds malachite green derivatives to produce red 
                     fluorescence"
                     /translation="QAVVTQEPSVTVSPGGTVILTCGSGTGAVTSGHYANWFQQKPGQA
                     PRALIFDTDKKYSWTPGRFSGSLLGAKAALTISDAQPEDEAEYYCSLSDVDGYLFGGGT
                     QLTVLSGGGGSGGGGSGGGGSGGGGSQAVVTQEPSVTVSPGGTVILTCGSGTGAVTSGH
                     YANWFQQKPGQAPRALIFDTDKKYSWTPGRFSGSLLGAKAALTISDAQPEDEAEYYCSL
                     SDVDGYLFGGGTQLTVLS"
     CDS             4272..4301
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             4302..4331
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     primer_bind     complement(4385..4401)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4386..4402)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     terminator      4404..4651
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(4420..4438)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     primer_bind     complement(4669..4688)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4670..4688)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4698..4715)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4698..4714)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4707..4729)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      4855..5310
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4942..4961)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5152..5173
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        5611..6018
                     /gene="S. cerevisiae LEU2"
                     /label=LEU2 promoter
     CDS             6019..7113
                     /codon_start=1
                     /gene="S. cerevisiae LEU2"
                     /product="3-isopropylmalate dehydrogenase, required for 
                     leucine biosynthesis"
                     /label=LEU2
                     /note="yeast auxotrophic marker"
                     /translation="MSAPKKIVVLPGDHVGQEITAEAIKVLKAISDVRSNVKFDFENHL
                     IGGAAIDATGVPLPDEALEASKKADAVLLGAVGGPKWGTGSVRPEQGLLKIRKELQLYA
                     NLRPCNFASDSLLDLSPIKPQFAKGTDFVVVRELVGGIYFGKRKEDDGDGVAWDSEQYT
                     VPEVQRITRMAAFMALQHEPPLPIWSLDKANVLASSRLWRKTVEETIKNEFPTLKVQHQ
                     LIDSAAMILVKNPTHLNGIIITSNMFGDIISDEASVIPGSLGLLPSASLASLPDKNTAF
                     GLYEPCHGSAPDLPKNKVNPIATILSAAMMLKLSLNLPEEGKAIEDAVKKVLDAGIRTG
                     DLGGSNSTTEVGDAVAEEVKKILA"
ORIGIN
        1 aaaaaaaaaa aatacaacaa ataaaaaaca ctcaatgacc tgaccatttg atggagttta
       61 agtcaatacc ttcttgaacc atttcccata atggtgaaag ttccctcaag aattttactc
      121 tgtcagaaac ggccttaacg acgtagtcga tatggtgcac tctcagtaca atctgctctg
      181 atgccgcata gttaagccag ccccgacacc cgccaacacc cgctgacgcg ccctgacggg
      241 cttgtctgct cccggcatcc gcttacagac aagctgtgac cgtctccggg agctgcatgt
      301 gtcagaggtt ttcaccgtca tcaccgaaac gcgcgagacg aaagggcctc gtgatacgcc
      361 tatttttata ggttaatgtc atgataataa tggtttctta ggacggatcg cttgcctgta
      421 acttacacgc gcctcgtatc ttttaatgat ggaataattt gggaatttac tctgtgttta
      481 tttattttta tgttttgtat ttggatttta gaaagtaaat aaagaaggta gaagagttac
      541 ggaatgaaga aaaaaaaata aacaaaggtt taaaaaattt caacaaaaag cgtactttac
      601 atatatattt attagacaag aaaagcagat taaatagata tacattcgat taacgataag
      661 taaaatgtaa aatcacagga ttttcgtgtg tggtcttcta cacagacaag atgaaacaat
      721 tcggcattaa tacctgagag caggaagagc aagataaaag gtagtatttg ttggcgatcc
      781 ccctagagtc ttttacatct tcggaaaaca aaaactattt tttctttaat ttcttttttt
      841 actttctatt tttaatttat atatttatat taaaaaattt aaattataat tatttttata
      901 gcacgtgatg aaaaggaccc aggtggcact tttcggggaa atgtgcgcgg aacccctatt
      961 tgtttatttt tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa
     1021 atgcttcaat aatattgaaa aaggaagagt atgagtattc aacatttccg tgtcgccctt
     1081 attccctttt ttgcggcatt ttgccttcct gtttttgctc acccagaaac gctggtgaaa
     1141 gtaaaagatg ctgaagatca gttgggtgca cgagtgggtt acatcgaact ggatctcaac
     1201 agcggtaaga tccttgagag ttttcgcccc gaagaacgtt ttccaatgat gagcactttt
     1261 aaagttctgc tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga gcaactcggt
     1321 cgccgcatac actattctca gaatgacttg gttgagtact caccagtcac agaaaagcat
     1381 cttacggatg gcatgacagt aagagaatta tgcagtgctg ccataaccat gagtgataac
     1441 actgcggcca acttacttct gacaacgatc ggaggaccga aggagctaac cgcttttttg
     1501 cacaacatgg gggatcatgt aactcgcctt gatcgttggg aaccggagct gaatgaagcc
     1561 ataccaaacg acgagcgtga caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa
     1621 ctattaactg gcgaactact tactctagct tcccggcaac aattaataga ctggatggag
     1681 gcggataaag ttgcaggacc acttctgcgc tcggcccttc cggctggctg gtttattgct
     1741 gataaatctg gagccggtga gcgtgggtct cgcggtatca ttgcagcact ggggccagat
     1801 ggtaagccct cccgtatcgt agttatctac acgacgggga gtcaggcaac tatggatgaa
     1861 cgaaatagac agatcgctga gataggtgcc tcactgatta agcattggta actgtcagac
     1921 caagtttact catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc
     1981 taggtgaaga tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc
     2041 cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg
     2101 cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg
     2161 gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca
     2221 aatactgttc ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg
     2281 cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg
     2341 tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga
     2401 acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac
     2461 ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat
     2521 ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc
     2581 tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga
     2641 tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc
     2701 ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg
     2761 gataaccgta ttaccgcctt tgagtgagct gataccgctc gccgcagccg aacgaccgag
     2821 cgcagcgagt cagtgagcga ggaagcggaa gagcgcccaa tacgcaaacc gcctctcccc
     2881 gcgcgttggc cgattcatta atgcagctgg cacgacaggt ttcccgactg gaaagcgggc
     2941 agtgagcgca acgcaattaa tgtgagttag ctcactcatt aggcacccca ggctttacac
     3001 tttatgcttc cggctcgtat gttgtgtgga attgtgagcg gataacaatt tcacacagga
     3061 aacagctatg accatgatta cgccaagcgc gcaattaacc ctcactaaag ggaacaaaag
     3121 ctggagctca tagcttcaaa atgtttctac tcctttttta ctcttccaga ttttctcgga
     3181 ctccgcgcat cgccgtacca cttcaaaaca cccaagcaca gcatactaaa tttcccctct
     3241 ttcttcctct agggtgtcgt taattacccg tactaaaggt ttggaaaaga aaaaagagac
     3301 cgcctcgttt ctttttcttc gtcgaaaaag gcaataaaaa tttttatcac gtttcttttt
     3361 cttgaaaatt ttttttttga tttttttctc tttcgatgac ctcccattga tatttaagtt
     3421 aataaacggt cttcaatttc tcaagtttca gtttcatttt tcttgttcta ttacaacttt
     3481 ttttacttct tgctcattag aaagaaagca tagcaatcta atctaagttt tctagaacta
     3541 gtggatccat gcaggccgtc gttacccaag aacctagtgt taccgttagc ccaggtggta
     3601 ctgttatact tacttgtgga agtggtacgg gtgccgtcac atctggtcat tatgcaaatt
     3661 ggtttcaaca aaaaccagga caagctccaa gagctttgat ttttgatact gataagaagt
     3721 attcttggac cccaggtaga ttttctggat ctttgctggg agcaaaggca gctttgacaa
     3781 tatcagatgc tcagcctgag gacgaagccg agtattactg ttctcttagc gacgtggatg
     3841 gctacttgtt tggcggtgga acacaactga cggttctgtc cggtggtggc ggctctggtg
     3901 gcggtggcag cggcggtggt ggttccggag gcggcggttc tcaggctgtg gtgactcagg
     3961 agccgtcagt gactgtgtcc ccaggaggga cagtcattct cacttgtggc tccggcactg
     4021 gagctgtcac cagtggtcat tatgccaact ggttccagca gaagcctggc caagccccca
     4081 gggcacttat atttgacacc gacaagaagt attcctggac ccctggccga ttctcaggct
     4141 ccctccttgg ggccaaggct gccctgacca tctcggatgc gcagcctgaa gatgaggctg
     4201 agtattactg ttcgctctcc gacgttgacg gttatctgtt cggaggaggc acccagctga
     4261 ccgtcctctc cgagcagaag ctgatcagcg aggaggacct ggagcagaag ctgatcagcg
     4321 aggaggacct ggcgggtgcg ggtgcgggtc gccccgggct gcaggaattc gatatcaagc
     4381 ttatcgatac cgtcgacctc gagtcatgta attagttatg tcacgcttac attcacgccc
     4441 tccccccaca tccgctctaa ccgaaaagga aggagttaga caacctgaag tctaggtccc
     4501 tatttatttt tttatagtta tgttagtatt aagaacgtta tttatatttc aaatttttct
     4561 tttttttctg tacagacgcg tgtacgcatg taacattata ctgaaaacct tgcttgagaa
     4621 ggttttggga cgctcgaagg ctttaatttg cggccggtac ccaattcgcc ctatagtgag
     4681 tcgtattacg cgcgctcact ggccgtcgtt ttacaacgtc gtgactggga aaaccctggc
     4741 gttacccaac ttaatcgcct tgcagcacat ccccctttcg ccagctggcg taatagcgaa
     4801 gaggcccgca ccgatcgccc ttcccaacag ttgcgcagcc tgaatggcga atggacgcgc
     4861 cctgtagcgg cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg accgctacac
     4921 ttgccagcgc cctagcgccc gctcctttcg ctttcttccc ttcctttctc gccacgttcg
     4981 ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt agggttccga tttagtgctt
     5041 tacggcacct cgaccccaaa aaacttgatt agggtgatgg ttcacgtagt gggccatcgc
     5101 cctgatagac ggtttttcgc cctttgacgt tggagtccac gttctttaat agtggactct
     5161 tgttccaaac tggaacaaca ctcaacccta tctcggtcta ttcttttgat ttataaggga
     5221 ttttgccgat ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga
     5281 attttaacaa aatattaacg cttacaattt cctgatgcgg tattttctcc ttacgcatct
     5341 gtgcggtatt tcacaccgca tatcgaccgg tcgaggagaa cttctagtat atctacatac
     5401 ctaatattat tgccttatta aaaatggaat cccaacaatt acatcaaaat ccacattctc
     5461 ttcaaaatca attgtcctgt acttccttgt tcatgtgtgt tcaaaaacgt tatatttata
     5521 ggataattat actctatttc tcaacaagta attggttgtt tggccgagcg gtctaaggcg
     5581 cctgattcaa gaaatatctt gaccgcagtt aactgtggga atactcaggt atcgtaagat
     5641 gcaagagttc gaatctctta gcaaccatta tttttttcct caacataacg agaacacaca
     5701 ggggcgctat cgcacagaat caaattcgat gactggaaat tttttgttaa tttcagaggt
     5761 cgcctgacgc atataccttt ttcaactgaa aaattgggag aaaaaggaaa ggtgagagcg
     5821 ccggaaccgg cttttcatat agaatagaga agcgttcatg actaaatgct tgcatcacaa
     5881 tacttgaagt tgacaatatt atttaaggac ctattgtttt ttccaatagg tggttagcaa
     5941 tcgtcttact ttctaacttt tcttaccttt tacatttcag caatatatat atatatattt
     6001 caaggatata ccattctaat gtctgcccct aagaagatcg tcgttttgcc aggtgaccac
     6061 gttggtcaag aaatcacagc cgaagccatt aaggttctta aagctatttc tgatgttcgt
     6121 tccaatgtca agttcgattt cgaaaatcat ttaattggtg gtgctgctat cgatgctaca
     6181 ggtgttccac ttccagatga ggcgctggaa gcctccaaga aggctgatgc cgttttgtta
     6241 ggtgctgtgg gtggtcctaa atggggtacc ggtagtgtta gacctgaaca aggtttacta
     6301 aaaatccgta aagaacttca attgtacgcc aacttaagac catgtaactt tgcatccgac
     6361 tctcttttag acttatctcc aatcaagcca caatttgcta aaggtactga cttcgttgtt
     6421 gtcagagaat tagtgggagg tatttacttt ggtaagagaa aggaagacga tggtgatggt
     6481 gtcgcttggg atagtgaaca atacaccgtt ccagaagtgc aaagaatcac aagaatggcc
     6541 gctttcatgg ccctacaaca tgagccacca ttgcctattt ggtccttgga taaagctaat
     6601 gttttggcct cttcaagatt atggagaaaa actgtggagg aaaccatcaa gaacgaattc
     6661 cctacattga aggttcaaca tcaattgatt gattctgccg ccatgatcct agttaagaac
     6721 ccaacccacc taaatggtat tataatcacc agcaacatgt ttggtgatat catctccgat
     6781 gaagcctccg ttatcccagg ttccttgggt ttgttgccat ctgcgtcctt ggcctctttg
     6841 ccagacaaga acaccgcatt tggtttgtac gaaccatgcc acggttctgc tccagatttg
     6901 ccaaagaata aggtcaaccc tatcgccact atcttgtctg ctgcaatgat gttgaaattg
     6961 tcattgaact tgcctgaaga aggtaaggcc attgaagatg cagttaaaaa ggttttggat
     7021 gcaggtatca gaactggtga tttaggtggt tccaacagta ccaccgaagt cggtgatgct
     7081 gtcgccgaag aagttaagaa aatccttgct taaaaagatt ctcttttttt atgatatttg
     7141 tacataaact ttataaatga aattcataat agaaacgaca cgaaattaca aaatggaata
     7201 tgttcatagg gtagacgaaa ctatatacgc aatctacata catttatcaa gaaggagaaa
     7261 aaggaggatg taaaggaata caggtaagca aattgatact aatggctcaa cgtgataagg
     7321 aaaaagaatt gcactttaac attaatattg acaaggagga gggcaccaca caaaaagtta
     7381 ggtgtaacag aaaatcatga aactatgatt cctaatttat atattggagg attttctcta
//
