LOCUS       sequence-407618-        6557 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian Expression of 3M-Qt-FLAG EMcapsulins.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6557)
  AUTHORS   Sigmund F, Berezin O, Beliakova S, Magerl B, Drawitsch M, Piovesan 
            A, Goncalves F, Bodea SV, Winkler S, Bousraou Z, Grosshauser M, 
            Samara E, Pujol-Marti J, Schadler S, So C, Irsen S, Walch A, Kofler 
            F, Piraud M, Kornfeld J, Briggman K, Westmeyer GG
  TITLE     Genetically encoded barcodes for correlative volume electron 
            microscopy.
  JOURNAL   Nat Biotechnol. 2023 Apr 17. doi: 10.1038/s41587-023-01713-y.
  PUBMED    37069313
REFERENCE   2  (bases 1 to 6557)
  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: "Nat 
            Biotechnol. 2023 Apr 17. doi: 10.1038/s41587-023-01713-y."
COMMENT     Sequence Label: 3M-Qt-FLAG
FEATURES             Location/Qualifiers
     source          1..6557
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      958..967
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2494..2517
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     primer_bind     complement(2565..2582)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2571..2795
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2841..3269
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2928..2947)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3138..3159
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3278..3298)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3283..3612
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3463..3598
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3525..3544
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        3660..3707
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             3726..4100
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     polyA_signal    4230..4351
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4267..4286)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4321..4340
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4400..4416)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4400..4416)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4413..4435)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4424..4440
                     /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)."
     promoter        complement(join(4448..4454,4455..4472,4473..4478))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4493..4514
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4631..4648)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4802..5390)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4882..4901)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5561..6352,6353..6421))
                     /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     6184..6203
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6422..6526)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc
      901 gtttaaactt aagcttggta ccgagctcgg atccactagt ccagtgtggt ggaattcgcc
      961 accatggacc ctgagacatg cccttgtcca acaggcggca gctgcacctg tagcgacaag
     1021 tgcaagtgca aaggctgcaa gtgtaccaac tgcaagaagt cctgctgcag ctgctgtcct
     1081 gccggctgtg aaaagtgcgc caaggactgc gtgtgcaagg gcgaagaagg cgccaaagcc
     1141 gaggccgaga agtgcagctg ttgtcagggc ggcaccggtg gcagcaccag caccacactg
     1201 gtcaagtgcg cctgcgagcc ttgcctgtgt aatgtggacc ctagcaaggc catcgaccgg
     1261 aacggcctgt actactgttc tgaggcctgt gccgatggcc acaccggcgg atctaaaggc
     1321 tgtggacaca ccggctgcaa ctgtcacggc tccgcgggtg gctgcgacga taagtgcgga
     1381 tgtgctgtgc cttgtcctgg cggcaccggc tgtagatgta caagcgctag aagcggagcc
     1441 gccgctggcg aacatacaac atgtggatgt ggcgagcact gcggctgcaa tccttgtgcc
     1501 tgtggcagag agggcacacc ttctggcaga gccaacagaa gggccaactg ttcttgtggc
     1561 gccgcttgca attgtgcctc ttgcggctct gctacagccc ctggaggtgg tggaagtggg
     1621 ggaggatcgg gtggatcagc gaacaagagc cagctgtacc ctgacagccc tctgaccgac
     1681 caggacttca accagctgga ccagaccgtg attgaagccg ccagaaggca gctcgtcggc
     1741 agaagattca tcgagctgta cggccctctc ggcagaggca tgcagagcgt gttcaacgac
     1801 atcttcatgg aaagccacga ggccaagatg gacttccagg gcagcttcga caccgaggtg
     1861 gaaagcagca gaagagtgaa ctacacaatc cccatgctgt acaaggactt cgtgctgtac
     1921 tggcgcgacc tggaacagag caaggccctg gacatcccca tcgatttttc cgtggccgcc
     1981 aacgccgcta gagatgtggc ctttctggaa gatcagatga tctttcacgg cagcaaagag
     2041 ttcgacatcc ccggcctgat gaacgtgaag ggcagactga cacacctgat cggcaattgg
     2101 tacgagagcg gcaacgcctt ccaggacatc gtggaagccc ggaacaagct gctggaaatg
     2161 aaccacaacg gcccctacgc tctggtgctg agccctgagc tgtatagcct gctgcacaga
     2221 gtgcacaagg acaccaacgt cctggaaatc gagcacgtgc gcgagctgat taccgccggt
     2281 gtctttcaga gccctgtgct gaagggaaag tccggcgtga tcgtgaacac cggcagaaac
     2341 aacctggacc tggccatcag cgaggacttc gagacagcct atctgggcga agagggcatg
     2401 aatcacccct tccgggtgta cgagacagtg gtgctgcgga ttaagaggcc tgccgccatc
     2461 tgcaccctga tcgatcctga agaaggctcc ggcgactaca aggacgacga cgacaaatga
     2521 gcggccgctc gagtctagag ggcccgttta aacccgctga tcagcctcga ctgtgccttc
     2581 tagttgccag ccatctgttg tttgcccctc ccccgtgcct tccttgaccc tggaaggtgc
     2641 cactcccact gtcctttcct aataaaatga ggaaattgca tcgcattgtc tgagtaggtg
     2701 tcattctatt ctggggggtg gggtggggca ggacagcaag ggggaggatt gggaagacaa
     2761 tagcaggcat gctggggatg cggtgggctc tatggcttct gaggcggaaa gaaccagctg
     2821 gggctctagg gggtatcccc acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt
     2881 ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt
     2941 cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcgggggct
     3001 ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgattaggg
     3061 tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga
     3121 gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca accctatctc
     3181 ggtctattct tttgatttat aagggatttt gccgatttcg gcctattggt taaaaaatga
     3241 gctgatttaa caaaaattta acgcgaatta attctgtgga atgtgtgtca gttagggtgt
     3301 ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca
     3361 gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat
     3421 ctcaattagt cagcaaccat agtcccgccc ctaactccgc ccatcccgcc cctaactccg
     3481 cccagttccg cccattctcc gccccatggc tgactaattt tttttattta tgcagaggcc
     3541 gaggccgcct ctgcctctga gctattccag aagtagtgag gaggcttttt tggaggccta
     3601 ggcttttgca aaaagctccc gggagcttgt atatccattt tcggatctga tcagcacgtg
     3661 ttgacaatta atcatcggca tagtatatcg gcatagtata atacgacaag gtgaggaact
     3721 aaaccatggc caagttgacc agtgccgttc cggtgctcac cgcgcgcgac gtcgccggag
     3781 cggtcgagtt ctggaccgac cggctcgggt tctcccggga cttcgtggag gacgacttcg
     3841 ccggtgtggt ccgggacgac gtgaccctgt tcatcagcgc ggtccaggac caggtggtgc
     3901 cggacaacac cctggcctgg gtgtgggtgc gcggcctgga cgagctgtac gccgagtggt
     3961 cggaggtcgt gtccacgaac ttccgggacg cctccgggcc ggccatgacc gagatcggcg
     4021 agcagccgtg ggggcgggag ttcgccctgc gcgacccggc cggcaactgc gtgcacttcg
     4081 tggccgagga gcaggactga cacgtgctac gagatttcga ttccaccgcc gccttctatg
     4141 aaaggttggg cttcggaatc gttttccggg acgccggctg gatgatcctc cagcgcgggg
     4201 atctcatgct ggagttcttc gcccacccca acttgtttat tgcagcttat aatggttaca
     4261 aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt
     4321 gtggtttgtc caaactcatc aatgtatctt atcatgtctg tataccgtcg acctctagct
     4381 agagcttggc gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa
     4441 ttccacacaa catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga
     4501 gctaactcac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt
     4561 gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct
     4621 cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat
     4681 cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga
     4741 acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt
     4801 ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt
     4861 ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc
     4921 gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa
     4981 gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct
     5041 ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta
     5101 actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg
     5161 gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc
     5221 ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta
     5281 ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg
     5341 gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt
     5401 tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg
     5461 tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta
     5521 aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg
     5581 aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg
     5641 tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc
     5701 gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg
     5761 agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg
     5821 aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag
     5881 gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat
     5941 caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc
     6001 cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc
     6061 ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa
     6121 ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac
     6181 gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt
     6241 cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc
     6301 gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa
     6361 caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca
     6421 tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat
     6481 acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa
     6541 aagtgccacc tgacgtc
//
