LOCUS       sequence-275853-        6527 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses a regulatable secretory cargo for mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6527)
  AUTHORS   Casler JC, Zajac AL, Valbuena FM, Sparvoli D, Jeyifous O, Turkewitz 
            AP, Horne-Badovinac S, Green WN, Glick BS
  TITLE     ESCargo: a regulatable fluorescent secretory cargo for diverse model
            organisms.
  JOURNAL   Mol Biol Cell. 2020 Oct 28:mbcE20090591. doi: 
            10.1091/mbc.E20-09-0591.
  PUBMED    33112725
REFERENCE   2  (bases 1 to 6527)
  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. 2020 Oct 28:mbcE20090591. doi: 10.1091/mbc.E20-09-0591."
COMMENT     Sequence Label: pEF-pIgH-ESCargo(FTV,Crimson)
FEATURES             Location/Qualifiers
     source          1..6527
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        4..1166
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          235..1157
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1114..1134
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     regulatory      1174..1183
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     sig_peptide     join(1180..1224,1307..1318)
                     /codon_start=1
                     /product="modified signal sequence from the mouse 
                     immunoglobulin heavy chain variable region"
                     /label=IgH signal sequence
                     /translation="MGWSCIILFLVATATGAHS"
     CDS             join(1180..1225,1308..1318)
                     /codon_start=1
                     /product="signal sequence from the mouse immunoglobulin 
                     heavy chain variableregion"
                     /label=IgH signal sequence
                     /translation="MGWSCIILFLVATATGAHS"
     CDS             1337..2008
                     /codon_start=1
                     /product="far-red noncytotoxic tetrameric variant of DsRed 
                     fluorescent protein (Strack et al., 2009)"
                     /label=E2-Crimson
                     /note="mammalian codon-optimized"
                     /translation="DSTENVIKPFMRFKVHMEGSVNGHEFEIEGVGEGKPYEGTQTAKL
                     QVTKGGPLPFAWDILSPQFFYGSKAYIKHPADIPDYLKQSFPEGFKWERVMNFEDGGVV
                     TVTQDSSLQDGTLIYHVKFIGVNFPSDGPVMQKKTLGWEPSTERNYPRDGVLKGENHMA
                     LKLKGGGHYLCEFKSIYMAKKPVKLPGYHYVDYKLDITSHNEDYTVVEQYERAEARHHL
                     FQ"
     primer_bind     1732..1751
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             2021..2341
                     /codon_start=1
                     /product="destabilization domain that can be stabilized by 
                     Shield1 in the ProteoTuner(TM) system"
                     /label=DD
                     /note="L106P mutant of FKBP12"
                     /translation="GVQVETISPGDGRTFPKRGQTVVVHYTGMLEDGKKLDSSRDRNKP
                     FKFMLGKQEVIRGWEEGVAQMSVGQRAKLTISPDYAYGATGHPGVIPPHATLVFDVELL
                     KLE"
     primer_bind     complement(2365..2382)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2371..2595
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2641..3069
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2728..2747)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2938..2959
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3078..3098)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3083..3412
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3263..3398
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3325..3344
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3438..4232
                     /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="MVEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3492..3511)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4102..4121
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4406..4527
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4443..4462)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4497..4516
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4560..4577)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4731..5319)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4811..4830)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5490..6281,6282..6350))
                     /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     6113..6132
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6351..6455)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 tgaggctccg gtgcccgtca gtgggcagag cgcacatcgc ccacagtccc cgagaagttg
       61 gggggagggg tcggcaattg aaccggtgcc tagagaaggt ggcgcggggt aaactgggaa
      121 agtgatgtcg tgtactggct ccgccttttt cccgagggtg ggggagaacc gtatataagt
      181 gcagtagtcg ccgtgaacgt tctttttcgc aacgggtttg ccgccagaac acaggtaagt
      241 gccgtgtgtg gttcccgcgg gcctggcctc tttacgggtt atggcccttg cgtgccttga
      301 attacttcca cctggctcca gtacgtgatt cttgatcccg agctggagcc aggggcgggc
      361 cttgcgcttt aggagcccct tcgcctcgtg cttgagttga ggcctggcct gggcgctggg
      421 gccgccgcgt gcgaatctgg tggcaccttc gcgcctgtct cgctgctttc gataagtctc
      481 tagccattta aaatttttga tgacctgctg cgacgctttt tttctggcaa gatagtcttg
      541 taaatgcggg ccaggatctg cacactggta tttcggtttt tggggccgcg ggcggcgacg
      601 gggcccgtgc gtcccagcgc acatgttcgg cgaggcgggg cctgcgagcg cggccaccga
      661 gaatcggacg ggggtagtct caagctggcc ggcctgctct ggtgcctggc ctcgcgccgc
      721 cgtgtatcgc cccgccctgg gcggcaaggc tggcccggtc ggcaccagtt gcgtgagcgg
      781 aaagatggcc gcttcccggc cctgctccag ggggctcaaa atggaggacg cggcgctcgg
      841 gagagcgggc gggtgagtca cccacacaaa ggaaaggggc ctttccgtcc tcagccgtcg
      901 cttcatgtga ctccacggag taccgggcgc cgtccaggca cctcgattag ttctggagct
      961 tttggagtac gtcgtcttta ggttgggggg aggggtttta tgcgatggag tttccccaca
     1021 ctgagtgggt ggagactgaa gttaggccag cttggcactt gatgtaattc tccttggaat
     1081 ttgccctttt tgagtttgga tcttggttca ttctcaagcc tcagacagtg gttcaaagtt
     1141 tttttcttcc atttcaggtg tcgtgaacac gtggccacca tgggatggag ctgtatcatc
     1201 ctcttcttgg tagcaacagc tacaggtaag gggttaacag tagcaggctt gaggtctgga
     1261 catatatatg ggtgacaatg acatccactt tgcctttctc tccacaggcg cgcactcctt
     1321 taccgtgaac accaccgata gcactgagaa cgtcatcaag cccttcatgc gcttcaaggt
     1381 gcacatggag ggctccgtga acggccacga gttcgagatc gagggcgtgg gcgagggcaa
     1441 gccctacgag ggcacccaga ccgccaagct gcaagtgacc aagggcggcc ccctgccctt
     1501 cgcctgggac atcctgtccc cccagttctt ctacggctcc aaggcgtaca tcaagcaccc
     1561 cgccgacatc cccgactacc tcaagcagtc cttccccgag ggcttcaagt gggagcgcgt
     1621 gatgaacttc gaggacggcg gcgtggtgac cgtgacccag gactcctccc tgcaggacgg
     1681 caccctcatc taccacgtga agttcatcgg cgtgaacttc ccctccgacg gccccgtaat
     1741 gcagaagaag actctgggct gggagccctc cactgagcgc aactaccccc gcgacggcgt
     1801 gctgaagggc gagaaccaca tggcgctgaa gctgaagggc ggcggccact acctgtgtga
     1861 gttcaagtcc atctacatgg ccaagaagcc cgtgaagctg cccggctacc actacgtgga
     1921 ctacaagctc gacatcacct cccacaacga ggactacacc gtggtggagc agtacgagcg
     1981 cgccgaggcc cgccaccacc tgttccaggg aggatccaga ggagtgcagg tggaaaccat
     2041 ctccccagga gacgggcgca ccttccccaa gcgcggccag accgttgtgg tgcactacac
     2101 cgggatgctt gaagatggaa agaaactaga ttcctcccgg gacagaaaca agccctttaa
     2161 gtttatgcta ggcaagcagg aggtgatccg aggctgggaa gaaggggttg cccagatgag
     2221 tgtgggtcag agagccaaac tgactatatc tccagattat gcctatggtg ccactgggca
     2281 cccaggcgtt atcccaccac atgccactct cgtcttcgat gtggagcttc taaaactgga
     2341 atagtctaga agctcgctga tcagcctcga ctgtgccttc tagttgccag ccatctgttg
     2401 tttgcccctc ccccgtgcct tccttgaccc tggaaggtgc cactcccact gtcctttcct
     2461 aataaaatga ggaaattgca tcgcattgtc tgagtaggtg tcattctatt ctggggggtg
     2521 gggtggggca ggacagcaag ggggaggatt gggaagacaa tagcaggcat gctggggatg
     2581 gcccgggctc tatggcttct gaggcggaaa gaaccagctg gggctctagg gggtatcccc
     2641 acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg
     2701 ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca
     2761 cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta
     2821 gtgctttacg gcacctctcc cccaaaaaac ttgattaggg tgatggttca cgtagtgggc
     2881 catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg
     2941 gactcttgtt ccaaactgga acaacactca accctatctc ggtctattct tttgatttat
     3001 aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa caaaaattta
     3061 acgcgaatta attctgtgga atgtgtgtca gttagggtgt ggaaagtccc caggctcccc
     3121 agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccaggt gtggaaagtc
     3181 cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccat
     3241 agtcccgccc ctaactccgc ccatcccgcc cctaactccg cccagttccg cccattctcc
     3301 gcccctaggc tgactaattt tttttattta tgcagaggcc gaggccgcct ctgcctctga
     3361 gctattccag aagtagtgag gaggcttttt tggaggccta ggcttttgca aaaagctccc
     3421 cccccgggag gtccacaatg gttgaacaag atggattgca cgcaggttct ccggccgctt
     3481 gggtggagag gctattcggc tatgactggg cacaacagac aatcggctgc tctgatgccg
     3541 ccgtgttccg gctgtcagcg caggggcgcc cggttctttt tgtcaagacc gacctgtccg
     3601 gtgccctgaa tgaactccag gacgaggcag cgcggctatc gtggctggcc acgacgggcg
     3661 ttccttgcgc agctgtgctc gacgttgtca ctgaagcggg aagggactgg ctgctattgg
     3721 gcgaagtgcc ggggcaggat ctcctgtcat ctcaccttgc tcctgccgag aaagtatcca
     3781 tcatggctga tgcaatgcgg cggctgcata cgcttgatcc ggctacctgc ccattcgacc
     3841 accaagcgaa acatcgcatc gagcgagcac gtactcggat ggaagccggt cttgtcgatc
     3901 aggatgatct ggacgaagag catcaggggc tcgcgccagc cgaactgttc gccaggctca
     3961 aggcgcgtat gcccgacggc gaggatctcg tcgtgactca tggcgatgcc tgcttgccga
     4021 atatcatggt ggaaaatggc cgcttttctg gattcatcga ctgtggccgg ctgggtgtgg
     4081 cggaccgcta tcaggacata gcgttggcta cccgtgatat tgctgaagag cttggcggcg
     4141 aatgggctga ccgcttcctc gtgctttacg gtatcgccgc tcccgattcg cagcgcatcg
     4201 ccttctatcg ccttcttgac gagttcttct gagcgggact ctggggttcg aaatgaccga
     4261 ccaagcgacg cccaacctgc catcacgaga tttcgattcc accgccgcct tctatgaaag
     4321 gttgggcttc ggaatcgttt tccgggacgc cggctggatg atcctccagc gcggggatct
     4381 catgctggag ttcttcgccc accccaactt gtttattgca gcttataatg gttacaaata
     4441 aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg
     4501 tttgtccaaa ctcatcaatg tatcttatca tgtctgtata ccgtcgatct ttccgcttcc
     4561 tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca
     4621 aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca
     4681 aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg
     4741 ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg
     4801 acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt
     4861 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt
     4921 tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc
     4981 tgtatgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt
     5041 gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt
     5101 agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc
     5161 tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa
     5221 agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt
     5281 tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct
     5341 acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta
     5401 tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa
     5461 agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc
     5521 tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact
     5581 acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc
     5641 tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt
     5701 ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta
     5761 agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg
     5821 tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt
     5881 acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc
     5941 agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt
     6001 actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc
     6061 tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc
     6121 gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa
     6181 ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg ggcacccaac
     6241 tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa
     6301 aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt
     6361 tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa
     6421 tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct
     6481 gacgtcagat cgacggatcg ggagatcggt accgaattca agcttcg
//
