LOCUS       sequence-250355-        4413 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses Lyn11-iFAST in mammalian cells (inner plasma membrane 
            labeling).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4413)
  TITLE     iFAST-containing plasmids
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 4413)
  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            "Unpublished"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: pAG471 Lyn11-iFAST
FEATURES             Location/Qualifiers
     source          1..4413
                     /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"
     CDS             663..1037
                     /codon_start=1
                     /product="small protein tag, derived from photoactive 
                     yellow protein, that allows in vivo fluorescent labeling by
                     addition of a cell-permeant fluorogen (Plamont et al., 
                     2016)"
                     /label=Y-FAST
                     /translation="MEHVAFGSEDIENTLAKMDDGQLDGLAFGAIQLDGDGNILQYNAA
                     EGDITGRDPKQVIGKNFFKDVAPGTDSPEFYGKFKEGVASGNLNTMFEWMIPTSRGPTK
                     VKIHMKKALSGDSYWVFVKRV"
     CDS             1044..1073
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     polyA_signal    1201..1322
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1238..1257)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1292..1311
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(1329..1784)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1466..1487)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1678..1697
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        1811..1915
                     /gene="bla"
                     /label=AmpR promoter
     promoter        1917..2274
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        1917..2108
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(1940..1960)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2125..2260
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             2309..3103
                     /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             2309..3103
                     /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(2363..2382)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2973..2992
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(3291..3310)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    3335..3382
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      3711..4299
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4200..4219
                     /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 ccggactcag atctatgggc tgcatcaagt ccaagggcaa ggactccgcc ggcggcggct
      661 ccatggagca tgttgccttt ggcagtgagg acatcgagaa cactctggcc aaaatggacg
      721 acggacaact ggatgggttg gcctttggcg caattcagct cgatggtgac gggaatatcc
      781 tgcagtacaa tgctgctgaa ggagacatca caggcagaga tcccaaacag gtgattggga
      841 agaacttctt caaggatgtt gcacctggaa cggattctcc cgagttttac ggcaaattca
      901 aggaaggcgt agcgtcaggg aatctgaaca ccatgttcga atggatgata ccgacaagca
      961 ggggaccaac caaggtcaag atacacatga agaaagccct ttccggtgac agctattggg
     1021 tctttgtgaa acgggtggga tccgaacaaa agcttatttc tgaagaggac ttgtaatagg
     1081 cggccgcgac tctagatcat aatcagccat accacatttg tagaggtttt acttgcttta
     1141 aaaaacctcc cacacctccc cctgaacctg aaacataaaa tgaatgcaat tgttgttgtt
     1201 aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca
     1261 aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct
     1321 taaggcgtaa attgtaagcg ttaatatttt gttaaaattc gcgttaaatt tttgttaaat
     1381 cagctcattt tttaaccaat aggccgaaat cggcaaaatc ccttataaat caaaagaata
     1441 gaccgagata gggttgagtg ttgttccagt ttggaacaag agtccactat taaagaacgt
     1501 ggactccaac gtcaaagggc gaaaaaccgt ctatcagggc gatggcccac tacgtgaacc
     1561 atcaccctaa tcaagttttt tggggtcgag gtgccgtaaa gcactaaatc ggaaccctaa
     1621 agggagcccc cgatttagag cttgacgggg aaagccggcg aacgtggcga gaaaggaagg
     1681 gaagaaagcg aaaggagcgg gcgctagggc gctggcaagt gtagcggtca cgctgcgcgt
     1741 aaccaccaca cccgccgcgc ttaatgcgcc gctacagggc gcgtcaggtg gcacttttcg
     1801 gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc
     1861 gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga agagtcctga
     1921 ggcggaaaga accagctgtg gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc
     1981 ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccag gtgtggaaag
     2041 tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc
     2101 atagtcccgc ccctaactcc gcccatcccg cccctaactc cgcccagttc cgcccattct
     2161 ccgccccatg gctgactaat tttttttatt tatgcagagg ccgaggccgc ctcggcctct
     2221 gagctattcc agaagtagtg aggaggcttt tttggaggcc taggcttttg caaagatcga
     2281 tcaagagaca ggatgaggat cgtttcgcat gattgaacaa gatggattgc acgcaggttc
     2341 tccggccgct tgggtggaga ggctattcgg ctatgactgg gcacaacaga caatcggctg
     2401 ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt ttgtcaagac
     2461 cgacctgtcc ggtgccctga atgaactgca agacgaggca gcgcggctat cgtggctggc
     2521 cacgacgggc gttccttgcg cagctgtgct cgacgttgtc actgaagcgg gaagggactg
     2581 gctgctattg ggcgaagtgc cggggcagga tctcctgtca tctcaccttg ctcctgccga
     2641 gaaagtatcc atcatggctg atgcaatgcg gcggctgcat acgcttgatc cggctacctg
     2701 cccattcgac caccaagcga aacatcgcat cgagcgagca cgtactcgga tggaagccgg
     2761 tcttgtcgat caggatgatc tggacgaaga gcatcagggg ctcgcgccag ccgaactgtt
     2821 cgccaggctc aaggcgagca tgcccgacgg cgaggatctc gtcgtgaccc atggcgatgc
     2881 ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct ggattcatcg actgtggccg
     2941 gctgggtgtg gcggaccgct atcaggacat agcgttggct acccgtgata ttgctgaaga
     3001 gcttggcggc gaatgggctg accgcttcct cgtgctttac ggtatcgccg ctcccgattc
     3061 gcagcgcatc gccttctatc gccttcttga cgagttcttc tgagcgggac tctggggttc
     3121 gaaatgaccg accaagcgac gcccaacctg ccatcacgag atttcgattc caccgccgcc
     3181 ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg ccggctggat gatcctccag
     3241 cgcggggatc tcatgctgga gttcttcgcc caccctaggg ggaggctaac tgaaacacgg
     3301 aaggagacaa taccggaagg aacccgcgct atgacggcaa taaaaagaca gaataaaacg
     3361 cacggtgttg ggtcgtttgt tcataaacgc ggggttcggt cccagggctg gcactctgtc
     3421 gataccccac cgagacccca ttggggccaa tacgcccgcg tttcttcctt ttccccaccc
     3481 caccccccaa gttcgggtga aggcccaggg ctcgcagcca acgtcggggc ggcaggccct
     3541 gccatagcct caggttactc atatatactt tagattgatt taaaacttca tttttaattt
     3601 aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag
     3661 ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct
     3721 ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt
     3781 tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg
     3841 cagataccaa atactgtcct tctagtgtag ccgtagttag gccaccactt caagaactct
     3901 gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc
     3961 gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg
     4021 tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa
     4081 ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg
     4141 gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg
     4201 ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga
     4261 tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt
     4321 ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct
     4381 gattctgtgg ataaccgtat taccgccatg cat
//
