LOCUS       sequence-409219-        6113 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian Expression of 1M-Mx-FLAG EMcapsulins.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6113)
  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 6113)
  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: 1M-Mx-FLAG
FEATURES             Location/Qualifiers
     source          1..6113
                     /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             2050..2073
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     primer_bind     complement(2121..2138)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2127..2351
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2397..2825
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2484..2503)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2694..2715
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2834..2854)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2839..3168
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3019..3154
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3081..3100
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        3216..3263
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             3282..3656
                     /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    3786..3907
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3823..3842)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3877..3896
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3956..3972)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3956..3972)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3969..3991)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3980..3996
                     /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(4004..4010,4011..4028,4029..4034))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4049..4070
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4187..4204)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4358..4946)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4438..4457)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5117..5908,5909..5977))
                     /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     5740..5759
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5978..6082)
                     /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 agtgctcttg ttgtcagggc agcggcccgg gccctgactt cctgggccac
     1201 gccgagaacc ccctgagaga agaggaatgg gccagactga acgagacagt gatccaggtg
     1261 gccaggcgga gcctcgtggg cagaagaatc ctggacatct acggccctct gggcgctggc
     1321 gtgcagacag tgccttacga tgagttccag ggcgtgtccc ctggcgccgt ggatattgtg
     1381 ggcgagcagg aaaccgctat ggtgttcacc gacgccagaa agttcaagac catccctatt
     1441 atctacaagg acttcctgct gcactggcgg gacatcgagg ccgccagaac ccacaacatg
     1501 cccctggatg tgtctgccgc tgctggcgct gctgctctgt gtgcccagca ggaagatgag
     1561 ctgatctttt acggcgacgc caggctgggc tacgagggcc tgatgacagc taacggcaga
     1621 ctgaccgtgc ccctgggcga ttggacatct cctggcggcg gattccaggc catcgtggaa
     1681 gccaccagaa agctgaacga gcagggccac ttcggcccct acgctgtggt gctgagcccc
     1741 agactgtaca gccagctgca cagaatctac gaaaagaccg gcgtgctgga aatcgagaca
     1801 atcagacagc tggcctccga cggcgtgtac cagagcaaca gactgagagg cgagagcggc
     1861 gtggtggtgt ccaccggcag agaaaacatg gacctggccg tgtctatgga catggtggcc
     1921 gcttacctgg gcgcctccag aatgaaccac cccttcagag tgctggaagc cctgctgctg
     1981 agaatcaagc accccgacgc catctgcacc ctggaaggcg caggcgccac agagagaaga
     2041 gggtcaggag attacaaaga tgacgatgat aaatgagcgg ccgctcgagt ctagagggcc
     2101 cgtttaaacc cgctgatcag cctcgactgt gccttctagt tgccagccat ctgttgtttg
     2161 cccctccccc gtgccttcct tgaccctgga aggtgccact cccactgtcc tttcctaata
     2221 aaatgaggaa attgcatcgc attgtctgag taggtgtcat tctattctgg ggggtggggt
     2281 ggggcaggac agcaaggggg aggattggga agacaatagc aggcatgctg gggatgcggt
     2341 gggctctatg gcttctgagg cggaaagaac cagctggggc tctagggggt atccccacgc
     2401 gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac
     2461 acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt
     2521 cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc
     2581 tttacggcac ctcgacccca aaaaacttga ttagggtgat ggttcacgta gtgggccatc
     2641 gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact
     2701 cttgttccaa actggaacaa cactcaaccc tatctcggtc tattcttttg atttataagg
     2761 gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc
     2821 gaattaattc tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg ctccccagca
     2881 ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca
     2941 ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc aaccatagtc
     3001 ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca ttctccgccc
     3061 catggctgac taattttttt tatttatgca gaggccgagg ccgcctctgc ctctgagcta
     3121 ttccagaagt agtgaggagg cttttttgga ggcctaggct tttgcaaaaa gctcccggga
     3181 gcttgtatat ccattttcgg atctgatcag cacgtgttga caattaatca tcggcatagt
     3241 atatcggcat agtataatac gacaaggtga ggaactaaac catggccaag ttgaccagtg
     3301 ccgttccggt gctcaccgcg cgcgacgtcg ccggagcggt cgagttctgg accgaccggc
     3361 tcgggttctc ccgggacttc gtggaggacg acttcgccgg tgtggtccgg gacgacgtga
     3421 ccctgttcat cagcgcggtc caggaccagg tggtgccgga caacaccctg gcctgggtgt
     3481 gggtgcgcgg cctggacgag ctgtacgccg agtggtcgga ggtcgtgtcc acgaacttcc
     3541 gggacgcctc cgggccggcc atgaccgaga tcggcgagca gccgtggggg cgggagttcg
     3601 ccctgcgcga cccggccggc aactgcgtgc acttcgtggc cgaggagcag gactgacacg
     3661 tgctacgaga tttcgattcc accgccgcct tctatgaaag gttgggcttc ggaatcgttt
     3721 tccgggacgc cggctggatg atcctccagc gcggggatct catgctggag ttcttcgccc
     3781 accccaactt gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt
     3841 tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg
     3901 tatcttatca tgtctgtata ccgtcgacct ctagctagag cttggcgtaa tcatggtcat
     3961 agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa
     4021 gcataaagtg taaagcctgg ggtgcctaat gagtgagcta actcacatta attgcgttgc
     4081 gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc
     4141 aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact
     4201 cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac
     4261 ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa
     4321 aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg
     4381 acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa
     4441 gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc
     4501 ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac
     4561 gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac
     4621 cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg
     4681 taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt
     4741 atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa
     4801 cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct
     4861 cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga
     4921 ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg
     4981 ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct
     5041 tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt
     5101 aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc
     5161 tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg
     5221 gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag
     5281 atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt
     5341 tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag
     5401 ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt
     5461 ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca
     5521 tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg
     5581 ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat
     5641 ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta
     5701 tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca
     5761 gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct
     5821 taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat
     5881 cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa
     5941 agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt
     6001 gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa
     6061 ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctgac gtc
//
