LOCUS       sequence-275932-        6509 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses a regulatable secretory cargo (without an ER export 
            signal) for mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6509)
  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 6509)
  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*
FEATURES             Location/Qualifiers
     source          1..6509
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        50..1212
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          281..1203
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1160..1180
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     regulatory      1220..1229
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     sig_peptide     join(1226..1270,1353..1364)
                     /codon_start=1
                     /product="modified signal sequence from the mouse 
                     immunoglobulin heavy chain variable region"
                     /label=IgH signal sequence
                     /translation="MGWSCIILFLVATATGAHS"
     CDS             join(1226..1271,1354..1364)
                     /codon_start=1
                     /product="signal sequence from the mouse immunoglobulin 
                     heavy chain variableregion"
                     /label=IgH signal sequence
                     /translation="MGWSCIILFLVATATGAHS"
     CDS             1365..2036
                     /codon_start=1
                     /product="noncytotoxic tetrameric variant of DsRed 
                     fluorescent protein (Strack et al., 2008)"
                     /label=DsRed-Express2
                     /note="mammalian codon-optimized"
                     /translation="DSTENVIKPFMRFKVHMEGSVNGHEFEIEGEGEGKPYEGTQTAKL
                     QVTKGGPLPFAWDILSPQFQYGSKVYVKHPADIPDYKKLSFPEGFKWERVMNFEDGGVV
                     TVTQDSSLQDGTFIYHVKFIGVNFPSDGPVMQKKTLGWEPSTERLYPRDGVLKGEIHKA
                     LKLKGGGHYLVEFKSIYMAKKPVKLPGYYYVDSKLDITSHNEDYTVVEQYERAEARHHL
                     FQ"
     primer_bind     complement(1542..1562)
                     /label=DsRed1-N
                     /note="DsRed1, reverse primer"
     primer_bind     1760..1779
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     1954..1977
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     CDS             2049..2369
                     /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(2393..2410)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2399..2623
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2669..3097
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2756..2775)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2966..2987
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3106..3126)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3111..3440
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3291..3426
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3353..3372
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3466..4260
                     /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(3520..3539)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4130..4149
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4434..4555
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4471..4490)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4525..4544
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4588..4605)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4759..5347)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4839..4858)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5518..6309,6310..6378))
                     /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     6141..6160
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6379..6483)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 acgtcagatc gacggatcgg gagatcggta ccgaattcaa gcttcgtgag gctccggtgc
       61 ccgtcagtgg gcagagcgca catcgcccac agtccccgag aagttggggg gaggggtcgg
      121 caattgaacc ggtgcctaga gaaggtggcg cggggtaaac tgggaaagtg atgtcgtgta
      181 ctggctccgc ctttttcccg agggtggggg agaaccgtat ataagtgcag tagtcgccgt
      241 gaacgttctt tttcgcaacg ggtttgccgc cagaacacag gtaagtgccg tgtgtggttc
      301 ccgcgggcct ggcctcttta cgggttatgg cccttgcgtg ccttgaatta cttccacctg
      361 gctccagtac gtgattcttg atcccgagct ggagccaggg gcgggccttg cgctttagga
      421 gccccttcgc ctcgtgcttg agttgaggcc tggcctgggc gctggggccg ccgcgtgcga
      481 atctggtggc accttcgcgc ctgtctcgct gctttcgata agtctctagc catttaaaat
      541 ttttgatgac ctgctgcgac gctttttttc tggcaagata gtcttgtaaa tgcgggccag
      601 gatctgcaca ctggtatttc ggtttttggg gccgcgggcg gcgacggggc ccgtgcgtcc
      661 cagcgcacat gttcggcgag gcggggcctg cgagcgcggc caccgagaat cggacggggg
      721 tagtctcaag ctggccggcc tgctctggtg cctggcctcg cgccgccgtg tatcgccccg
      781 ccctgggcgg caaggctggc ccggtcggca ccagttgcgt gagcggaaag atggccgctt
      841 cccggccctg ctccaggggg ctcaaaatgg aggacgcggc gctcgggaga gcgggcgggt
      901 gagtcaccca cacaaaggaa aggggccttt ccgtcctcag ccgtcgcttc atgtgactcc
      961 acggagtacc gggcgccgtc caggcacctc gattagttct ggagcttttg gagtacgtcg
     1021 tctttaggtt ggggggaggg gttttatgcg atggagtttc cccacactga gtgggtggag
     1081 actgaagtta ggccagcttg gcacttgatg taattctcct tggaatttgc cctttttgag
     1141 tttggatctt ggttcattct caagcctcag acagtggttc aaagtttttt tcttccattt
     1201 caggtgtcgt gaacacgtgg ccaccatggg atggagctgt atcatcctct tcttggtagc
     1261 aacagctaca ggtaaggggt taacagtagc aggcttgagg tctggacata tatatgggtg
     1321 acaatgacat ccactttgcc tttctctcca caggcgcgca ctccgatagc actgagaacg
     1381 tcatcaagcc cttcatgcgc ttcaaggtgc acatggaggg ctcagtaaat ggtcatgagt
     1441 tcgagatcga gggcgagggc gagggcaagc cctacgaggg cacccagacc gccaagctgc
     1501 aggtgaccaa gggcggcccc ctgcccttcg cctgggacat cctgtccccc cagttccagt
     1561 acggctccaa ggtgtacgtg aagcaccccg ccgacatccc cgactacaag aagctgtcct
     1621 tccccgaggg cttcaagtgg gagcgcgtga tgaacttcga ggacggcggc gtggtgaccg
     1681 tgacccagga ctcctccctg caggacggca ccttcatcta ccacgtgaag ttcatcggcg
     1741 tgaacttccc ctccgacggc cccgtaatgc agaagaagac tctgggctgg gagccctcca
     1801 ccgagcgcct gtacccccgc gacggcgtgc tgaagggcga gatccacaag gcgctgaagc
     1861 tgaagggcgg cggccactac ctggtggagt tcaagtcaat ctacatggcc aagaagcccg
     1921 tgaagctgcc cggctactac tacgtggact ccaagctgga catcacctcc cacaacgagg
     1981 actacaccgt ggtggagcag tacgagcgcg ccgaggcccg ccaccacctg ttccagggag
     2041 gatccagagg agtgcaggtg gaaaccatct ccccaggaga cgggcgcacc ttccccaagc
     2101 gcggccagac cgttgtggtg cactacaccg ggatgcttga agatggaaag aaactagatt
     2161 cctcccggga cagaaacaag ccctttaagt ttatgctagg caagcaggag gtgatccgag
     2221 gctgggaaga aggggttgcc cagatgagtg tgggtcagag agccaaactg actatatctc
     2281 cagattatgc ctatggtgcc actgggcacc caggcgttat cccaccacat gccactctcg
     2341 tcttcgatgt ggagcttcta aaactggaat agtctagaag ctcgctgatc agcctcgact
     2401 gtgccttcta gttgccagcc atctgttgtt tgcccctccc ccgtgccttc cttgaccctg
     2461 gaaggtgcca ctcccactgt cctttcctaa taaaatgagg aaattgcatc gcattgtctg
     2521 agtaggtgtc attctattct ggggggtggg gtggggcagg acagcaaggg ggaggattgg
     2581 gaagacaata gcaggcatgc tggggatggc ccgggctcta tggcttctga ggcggaaaga
     2641 accagctggg gctctagggg gtatccccac gcgccctgta gcggcgcatt aagcgcggcg
     2701 ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca gcgccctagc gcccgctcct
     2761 ttcgctttct tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat
     2821 cgggggctcc ctttagggtt ccgatttagt gctttacggc acctctcccc caaaaaactt
     2881 gattagggtg atggttcacg tagtgggcca tcgccctgat agacggtttt tcgccctttg
     2941 acgttggagt ccacgttctt taatagtgga ctcttgttcc aaactggaac aacactcaac
     3001 cctatctcgg tctattcttt tgatttataa gggattttgc cgatttcggc ctattggtta
     3061 aaaaatgagc tgatttaaca aaaatttaac gcgaattaat tctgtggaat gtgtgtcagt
     3121 tagggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca
     3181 attagtcagc aaccaggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa
     3241 gcatgcatct caattagtca gcaaccatag tcccgcccct aactccgccc atcccgcccc
     3301 taactccgcc cagttccgcc cattctccgc ccctaggctg actaattttt tttatttatg
     3361 cagaggccga ggccgcctct gcctctgagc tattccagaa gtagtgagga ggcttttttg
     3421 gaggcctagg cttttgcaaa aagctccccc cccgggaggt ccacaatggt tgaacaagat
     3481 ggattgcacg caggttctcc ggccgcttgg gtggagaggc tattcggcta tgactgggca
     3541 caacagacaa tcggctgctc tgatgccgcc gtgttccggc tgtcagcgca ggggcgcccg
     3601 gttctttttg tcaagaccga cctgtccggt gccctgaatg aactccagga cgaggcagcg
     3661 cggctatcgt ggctggccac gacgggcgtt ccttgcgcag ctgtgctcga cgttgtcact
     3721 gaagcgggaa gggactggct gctattgggc gaagtgccgg ggcaggatct cctgtcatct
     3781 caccttgctc ctgccgagaa agtatccatc atggctgatg caatgcggcg gctgcatacg
     3841 cttgatccgg ctacctgccc attcgaccac caagcgaaac atcgcatcga gcgagcacgt
     3901 actcggatgg aagccggtct tgtcgatcag gatgatctgg acgaagagca tcaggggctc
     3961 gcgccagccg aactgttcgc caggctcaag gcgcgtatgc ccgacggcga ggatctcgtc
     4021 gtgactcatg gcgatgcctg cttgccgaat atcatggtgg aaaatggccg cttttctgga
     4081 ttcatcgact gtggccggct gggtgtggcg gaccgctatc aggacatagc gttggctacc
     4141 cgtgatattg ctgaagagct tggcggcgaa tgggctgacc gcttcctcgt gctttacggt
     4201 atcgccgctc ccgattcgca gcgcatcgcc ttctatcgcc ttcttgacga gttcttctga
     4261 gcgggactct ggggttcgaa atgaccgacc aagcgacgcc caacctgcca tcacgagatt
     4321 tcgattccac cgccgccttc tatgaaaggt tgggctttgg aatcgttttc cgggacgccg
     4381 gctggatgat cctccagcgc ggggatctca tgctggagtt cttcgcccac cccaacttgt
     4441 ttattgcagc ttataatggt tacaaataaa gcaatagcat cacaaatttc acaaataaag
     4501 catttttttc actgcattct agttgtggtt tgtccaaact catcaatgta tcttatcatg
     4561 tctgtatacc gtcgatcttt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg
     4621 gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg
     4681 ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa
     4741 ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg
     4801 acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc
     4861 tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc
     4921 ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc
     4981 ggtgtaggtc gttcgctcca agctgggctg tatgcacgaa ccccccgttc agcccgaccg
     5041 ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc
     5101 actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga
     5161 gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg gtatctgcgc
     5221 tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac
     5281 caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg
     5341 atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc
     5401 acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa
     5461 ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta
     5521 ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt
     5581 tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag
     5641 tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag caataaacca
     5701 gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc
     5761 tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt
     5821 tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag
     5881 ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt
     5941 tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat
     6001 ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt
     6061 gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc
     6121 ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat
     6181 cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag
     6241 ttcgatgtaa cccactcggg cacccaactg atcttcagca tcttttactt tcaccagcgt
     6301 ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg
     6361 gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta
     6421 ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc
     6481 gcgcacattt ccccgaaaag tgccacctg
//
