LOCUS       sequence-493000-        7352 bp    DNA     circular SYN 18-NOV-2025
DEFINITION  Expresses FAP-GFP-KDEL in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7352)
  AUTHORS   Moore WM, Brea RJ, Knittel C, Wrightsman E, Hui B, Lou J, Ancajas 
            CF, Best MD, Obara, CJ Devaraj NK, Budin I
  TITLE     Leaflet-specific phospholipid imaging using genetically encoded 
            proximity sensors
  JOURNAL   Nat Chem Biol (2025). https://doi.org/10.1038/s41589-025-02021-z.
  PUBMED    38746395
REFERENCE   2  (bases 1 to 7352)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Nov 18, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat Chem 
            Biol (2025). https://doi.org/10.1038/s41589-025-02021-z."
COMMENT     Sequence Label: pSBbi-FAP-GFP-KDEL
FEATURES             Location/Qualifiers
     source          1..7352
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     349..368
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     promoter        1000..2182
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     primer_bind     2130..2150
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     regulatory      2208..2217
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     sig_peptide     2214..2276
                     /codon_start=1
                     /product="leader sequence from mouse immunoglobulin kappa 
                     light chain"
                     /label=Ig-kappa leader
                     /translation="METDTLLLWVLLLWVPGSTGD"
     CDS             join(3195..3197,3198..3200,3201..3911)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(3195..3197,3198..3200,3201..3911)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(3240..3261)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3501..3520)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3848..3869
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(3952..3969)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3958..4182
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     promoter        4614..4718
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4719..4787,4788..5579)
                     /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(4937..4956)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5750..6338
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6239..6258
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6492..6509
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     6883..6902
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     polyA_signal    6901..6956
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     7002..7021
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     CDS             complement(join(7121..7352,1..368))
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(7204..7224)
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
ORIGIN
        1 cacgcgggct ccttgggccg gtgcggcgcc aggaggcctt ccatctgttg ctgcgcggcc
       61 agccgggaac cgctcaactc ggccatgcgc gggccgatct cggcgaacac cgcccccgct
      121 tcgacgctct ccggcgtggt ccagaccgcc accgcggcgc cgtcgtccgc gacccacacc
      181 ttgccgatgt cgagcccgac gcgcgtgagg aagagttctt gcagctcggt gacccgctcg
      241 atgtggcggt ccggatcgac ggtgtggcgc gtggcggggt agtcggcgaa cgcggcggcg
      301 agggtgcgta cggccctggg gacgtcgtcg cgggtggcga ggcgcaccgt gggcttgtac
      361 tcggtcatgg tggcctcagg tgcagaggtt tctacaggga aagagagaaa ggctacagtt
      421 agtttgctgt gatgatgcta agtgccgagg tcagatgtct atggattgta cacggaatcc
      481 caacgcagga accaaaaaac actcacccaa aaaaatggcg ctaaggccga gagaaaggcg
      541 aagttccgtc tacggctata taccttgaaa caggggcggc agaagtgacg caacgcggtt
      601 gcgcggacgg cttcctgtgt tttcattggt cgaagcgttc tgcgcgtgcg cagagattcc
      661 ccagggaagg cgggccctct tccgaatgtc caatccggac attcttccgg ttggacctcc
      721 gcggtcagcg ctgtgcccgg cgagagatca cgtggggcgc ggaggcggtg ctgctggggc
      781 acggccgtcc agcctcggcg gccatatttt tgaggggctg ttcatctcgt tcacacgctc
      841 tgtccgccat gtttgtgagt ggaagcgcca ttaccccttc aagcgactga aggctgcagg
      901 gcctctggtg gcccgcatgg ggagaccaga cccgccaggc ccgcctttcc gcactcagtc
      961 cgggcttact ttattttgtg agacagggtc tcgcctagag gctccggtgc ccgtcagtgg
     1021 gcagagcgca catcgcccac agtccccgag aagttggggg gaggggtcgg caattgaacc
     1081 ggtgcctaga gaaggtggcg cggggtaaac tgggaaagtg atgtcgtgta ctggctccgc
     1141 ctttttcccg agggtggggg agaaccgtat ataagtgcag tagtcgccgt gaacgttctt
     1201 tttcgcaacg ggtttgccgc cagaacacag gtaagtgccg tgtgtggttc ccgcgggcct
     1261 ggcctcttta cgggttatgg cccttgcgtg ccttgaatta cttccacgcc cctggctgca
     1321 gtacgtgatt cttgatcccg agcttcgggt tggaagtggg tgggagagtt cgaggccttg
     1381 cgcttaagga gccccttcgc ctcgtgcttg agttgaggcc tggcttgggc gctggggccg
     1441 ccgcgtgcga atctggtggc accttcgcgc ctgtctcgct gctttcgata agtctctagc
     1501 catttaaaat ttttgatgac ctgctgcgac gctttttttc tggcaagata gtcttgtaaa
     1561 tgcgggccaa gatctgcaca ctggtatttc ggtttttggg gccgcgggcg gcgacggggc
     1621 ccgtgcgtcc cagcgcacat gttcggcgag gcggggcctg cgagcgcggc caccgagaat
     1681 cggacggggg tagtctcaag ctggccggcc tgctctggtg cctggcctcg cgccgccgtg
     1741 tatcgccccg ccctgggcgg caaggctggc ccggtcggca ccagttgcgt gagcggaaag
     1801 atggccgctt cccggccctg ctgcagggag ctcaaaatgg aggacgcggc gctcgggaga
     1861 gcgggcgggt gagtcaccca cacaaaggaa aagggccttt ccgtcctcag ccgtcgcttc
     1921 atgtgactcc acggagtacc gggcgccgtc caggcacctc gattagttct cgagcttttg
     1981 gagtacgtcg tctttaggtt ggggggaggg gttttatgcg atggagtttc cccatactga
     2041 gtgggtggag actgaagtta ggccagcttg gcacttgatg taattctcct tggaatttgc
     2101 cctttttgag tttggatctt ggttcattct caagcctcag acagtggttc aaagtttttt
     2161 tcttccattt aaggtgtcgt gaaaactacc ccaagctggc ctctgaggcc accatggaga
     2221 cagacacact cctgctatgg gtactgctgc tctgggttcc aggttccact ggtgacgaat
     2281 tcggcctttc tggcctgcaa gtccagttgc aagaatctgg acctggtttg gttaaaccat
     2341 ctgagacact ttccctaacc tgcacagtca gtggcgcatc aatttcctcc tcgcactatt
     2401 attggggttg gattagacaa ccacctggaa agggcccaga gtggataggg tctatgtact
     2461 actcgggtag aacttactat aatcctgcct tgaagtcacg tgttaccata tcctctgata
     2521 agtcaaaaaa ccaattcttt ttaaaactca ctagcgttac cgctgctgat acagctgtct
     2581 attatgctgc ccgtgagggc ccaacgcatt attatgataa ttcaggccca atacctagcg
     2641 atgagtattt tcaacattgg ggccagggca ctttagttac agtttccggt ggtggcggct
     2701 ctggtggcgg tggcagcggc ggtggtggtt ccggaggcgg cggttctcag gtgcagctgc
     2761 aggagtcggg cccaggactg gtgaagcctt cggagaccct gtccctcacc tgcactgtct
     2821 ctggtgcctc catcagcagt agtcattact actggggctg gattcgccag cccccaggga
     2881 aggggcctga gtggattggg agtatgtatt atagtgggag aacgtactac aacccggccc
     2941 tcaagagtcg agtcaccata tcatcagaca agtcgaagaa ccagttcttc ttgaagttga
     3001 cctctgtaac cgccgcggac acggccgtgt attacgctgc gagggaggga cccacacatt
     3061 actatgataa tagtggtcca ataccttcgg atgagtattt ccagcactgg ggccagggta
     3121 ccctggtcac tgtctccgcg gccggtctgg cccatatggg aggcggtggg tctggtggcg
     3181 gtggcagcac cggtatggtg agcaagggcg aggagctgtt caccggggtg gtgcccatcc
     3241 tggtcgagct ggacggcgac gtaaacggcc acaagttcag cgtgtccggc gagggcgagg
     3301 gcgatgccac ctacggcaag ctgaccctga agttcatctg caccaccggc aagctgcccg
     3361 tgccctggcc caccctcgtg accaccctga cctacggcgt gcagtgcttc agccgctacc
     3421 ccgaccacat gaagcagcac gacttcttca agtccgccat gcccgaaggc tacgtccagg
     3481 agcgcaccat cttcttcaag gacgacggca actacaagac ccgcgccgag gtgaagttcg
     3541 agggcgacac cctggtgaac cgcatcgagc tgaagggcat cgacttcaag gaggacggca
     3601 acatcctggg gcacaagctg gagtacaact acaacagcca caacgtctat atcatggccg
     3661 acaagcagaa gaacggcatc aaggtgaact tcaagatccg ccacaacatc gaggacggca
     3721 gcgtgcagct cgccgaccac taccagcaga acacccccat cggcgacggc cccgtgctgc
     3781 tgcccgacaa ccactacctg agcacccagt ccgccctgag caaagacccc aacgagaagc
     3841 gcgatcacat ggtcctgctg gagttcgtga ccgccgccgg gatcactctc ggcatggacg
     3901 agctgtacaa gctcgagaag gatgaactgt aatctagagc tcgctgatca gcctcgactg
     3961 tgccttctag ttgccagcca tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg
     4021 aaggtgccac tcccactgtc ctttcctaat aaaatgagga aattgcatcg cattgtctga
     4081 gtaggtgtca ttctattctg gggggtgggg tggggcagga cagcaagggg gaggattggg
     4141 aagacaatag caggcatgct ggggatgcgg tgggctctat ggcttctgag gcggaaagaa
     4201 ccagctgggc ctactagata gcggacccct taccgaaaca tcgccgcatt ctgcagagga
     4261 gtcgaactta gtgtatgtaa acttctgacc cactggaatt gtgatacagt gaattataag
     4321 tgaaataatc tgtctgtaaa caattgttgg aaaaatgact tgtgtcatgc acaaagtaga
     4381 tgtcctaact gacttgccaa aactattgtt tgttaacaag aaatttgtgg agtagttgaa
     4441 aaacgagttt taatgactcc aacttaagtg tatgtaaact tccgacttca actgtatagg
     4501 gatccaattt taacaaaata ttaacgctta caatttcctg atgcggtatt ttctccttac
     4561 gcatctgtgc ggtatttcac accgcatcag gtggcacttt tcggggaaat gtgcgcggaa
     4621 cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac
     4681 cctgataaat gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg
     4741 tcgcccttat tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc
     4801 tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg
     4861 atctcaacag cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga
     4921 gcacttttaa agttctgcta tgtggcgcgg tattatcccg tattgacgcc gggcaagagc
     4981 aactcggtcg ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag
     5041 aaaagcatct tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga
     5101 gtgataacac tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg
     5161 cttttttgca caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga
     5221 atgaagccat accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt
     5281 tgcgcaaact attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact
     5341 ggatggaggc ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt
     5401 ttattgctga taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg
     5461 ggccagatgg taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta
     5521 tggatgaacg aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac
     5581 tgtcagacca agtttactca tatatacttt agattgattt aaaacttcat ttttaattta
     5641 aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt
     5701 tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt
     5761 tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt
     5821 gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc
     5881 agataccaaa tactgttctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg
     5941 tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg
     6001 ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt
     6061 cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac
     6121 tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg
     6181 acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg
     6241 gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat
     6301 ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt
     6361 tacggttcct ggccttttgc tggccttttg ctcacatgtt ctttcctgcg ttatcccctg
     6421 attctgtgga taaccgtatt accgcctttg agtgagctga taccgctcgc cgcagccgaa
     6481 cgaccgagcg cagcgagtca gtgagcgagg aagcggatcc ctatacagtt gaagtcggaa
     6541 gtttacatac accttagcca aatacattta aactcacttt ttcacaattc ctgacattta
     6601 atcctagtaa aaattccctg tcttaggtca gttaggatca ccactttatt ttaagaatgt
     6661 gaaatatcag aataatagta gagagaatga ttcatttcag cttttatttc tttcatcaca
     6721 ttcccagtgg gtcagaagtt tacatacact cgactcctct gcagaatgcg gcgatgtttc
     6781 ggtaaggggt cccgagtagc tagttcatgg cagccagcat atggcatatg ttgccaaact
     6841 ctaaaccaaa tactcattct gatgttttaa atgatttgcc ctcccatatg tccttccgag
     6901 tgagagacac aaaaaattcc aacacactat tgcaatgaaa ataaatttcc tttattagcc
     6961 agaagtcaga tgctcaaggg gcttcatgat gtccccataa tttttggcag agggaaaaag
     7021 atctcagtgg tatttgtgag ccagggcatt ggccacacca gccaccacct tctgataggc
     7081 agcctgcacc tgaggagtga attcacgaca ggccttcgaa tcaggcaccg ggcttgcggg
     7141 tcatgcacca ggtgcgcggt ccttcgggca cctcgacgtc ggcggtgacg gtgaagccga
     7201 gccgctcgta gaaggggagg ttgcggggcg cggaggtctc caggaaggcg ggcaccccgg
     7261 cgcgctcggc cgcctccact ccggggagca cgacggcgct gcccagaccc ttgccctggt
     7321 ggtcgggcga gacgccgacg gtggccagga ac
//
