LOCUS       sequence-409223-        8249 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of heterobifunctional SasG linkers (5G).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8249)
  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 8249)
  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: sfGFP-SasG(5G)-mCherry
FEATURES             Location/Qualifiers
     source          1..8249
                     /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"
     CDS             join(964..966,967..1677)
                     /codon_start=1
                     /product="GFP variant that folds robustly even when fused 
                     to poorly folded proteins (Pedelacq et al., 2006)"
                     /label=superfolder GFP
                     /note="mammalian codon-optimized"
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTISFKDDG
                     TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA
                     NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     primer_bind     complement(4257..4274)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4263..4487
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      4533..4961
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4620..4639)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4830..4851
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4970..4990)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4975..5304
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5155..5290
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     5217..5236
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        5352..5399
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             5418..5792
                     /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    5922..6043
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5959..5978)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6013..6032
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(6092..6108)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6092..6108)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6105..6127)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6116..6132
                     /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(6140..6146,6147..6164,6165..6170))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6185..6206
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6323..6340)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6494..7082)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6574..6593)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7253..8044,8045..8113))
                     /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     7876..7895
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8114..8218)
                     /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 accatgagca agggcgaaga actgtttacc ggcgtggtgc ccatcctggt ggaactggat
     1021 ggggatgtga acggccacaa gttcagcgtc agaggcgaag gcgaagggga tgccacaaac
     1081 ggcaagctga ccctgaagtt catctgcacc accggaaagc tgcccgtgcc ttggcctaca
     1141 ctggtcacca cactgacata cggcgtgcag tgcttcagca gataccccga ccatatgaag
     1201 cggcacgact tcttcaagag cgccatgcct gagggctacg tgcaagagag aaccatcagc
     1261 ttcaaggacg acggcaccta caagaccaga gccgaagtga agttcgaggg cgacaccctg
     1321 gtcaaccgga tcgagctgaa gggcatcgat ttcaaagagg acggcaacat cctgggccac
     1381 aagcttgagt acaacttcaa cagccacaac gtgtacatca ccgccgacaa gcagaagaac
     1441 ggcatcaagg ccaacttcaa gatccggcac aacgtggaag atggcagcgt gcagctggcc
     1501 gatcactacc agcagaacac acccatcgga gatggccctg tgctgctgcc cgataaccac
     1561 tacctgagca cacagagcgt gctgagcaag gaccccaacg agaagaggga ccacatggtg
     1621 ctgctggagt ttgtgacagc cgccggaatc acccacggca tggacgaact ttacaaaggc
     1681 ggcaccggtg gcagcgcccc taagacaatt accgagctgg aaaagaaggt ggaagagatc
     1741 cccttcaaga aagagcggaa gttcaacccc gatctggccc ctggcaccga gaaagtgaca
     1801 agagaaggcc agaagggcga gaaaaccatc accacaccta cactgaagaa ccctctgacc
     1861 ggcgtgatca tctccaaggg cgagcccaaa gaggaaatca caaaggaccc tatcaacgag
     1921 ctgaccgagt acggccccga gacaattgct cctggccaca gagatgagtt cgaccccaaa
     1981 ctgcccaccg gcgagaaaga agaagtgcct ggcaagcctg gcatcaagaa ccctgagaca
     2041 ggcgacgtcg tcagacctcc tgtggacagc gtgaccaaat acggccctgt gaagggcgac
     2101 agcatcgtgg aaaaagaaga gattccgttt gagaaagaga gaaagtttaa tcccgacctc
     2161 gctccaggga cagaaaaagt cactcgcgag ggccagaaag gggaaaagac aatcacaacc
     2221 ccaacgctga aaaatcccct gaccggggag attatcagca aaggcgagag caaagaagaa
     2281 attacgaaag atcccatcaa tgaactcaca gagtacgggc ccgagacaat tgctcctggc
     2341 cacagagatg agttcgaccc caaactgccc accggcgaga aagaagaagt gcctggcaag
     2401 cctggcatca agaaccctga gacaggcgac gtcgtcagac ctcctgtgga cagcgtgacc
     2461 aaatacggcc ctgtgaaggg cgacagcatc gtggaaaaag aagagattcc gtttgagaaa
     2521 gagagaaagt ttaatcccga cctcgctcca gggacagaaa aagtcactcg cgagggccag
     2581 aaaggggaaa agacaatcac aaccccaacg ctgaaaaatc ccctgaccgg ggagattatc
     2641 agcaaaggcg agagcaaaga agaaattacg aaagatccca tcaatgaact cacagagtac
     2701 gggcccgaga caattgctcc tggccacaga gatgagttcg accccaaact gcccaccggc
     2761 gagaaagaag aagtgcctgg caagcctggc atcaagaacc ctgagacagg cgacgtcgtc
     2821 agacctcctg tggacagcgt gaccaaatac ggccctgtga agggcgacag catcgtggaa
     2881 aaagaagaga ttccgtttga gaaagagaga aagtttaatc ccgacctcgc tccagggaca
     2941 gaaaaagtca ctcgcgaggg ccagaaaggg gaaaagacaa tcacaacccc aacgctgaaa
     3001 aatcccctga ccggggagat tatcagcaaa ggcgagagca aagaagaaat tacgaaagat
     3061 cccatcaatg aactcacaga gtacgggccc gagacaattg ctcctggcca cagagatgag
     3121 ttcgacccca aactgcccac cggcgagaaa gaagaagtgc ctggcaagcc tggcatcaag
     3181 aaccctgaga caggcgacgt cgtcagacct cctgtggaca gcgtgaccaa atacggccct
     3241 gtgaagggcg acagcatcgt ggaaaaagaa gagattccgt ttgagaaaga gagaaagttt
     3301 aatcccgacc tcgctccagg gacagaaaaa gtcactcgcg agggccagaa aggggaaaag
     3361 acaatcacaa ccccaacgct gaaaaatccc ctgaccgggg agattatcag caaaggcgag
     3421 agcaaagaag aaattacgaa agatcccatc aatgaactca cagagtacgg gcctgagacg
     3481 ggctccggag ccacaggatc tgtgtccaaa ggcgaagagg acaacatggc catcatcaaa
     3541 gaatttatgc ggttcaaggt gcacatggaa ggctccgtga atggccacga gtttgagatt
     3601 gaaggcgaag gcgagggcag accctatgag ggaacacaga ccgccaagct gaaagtgacc
     3661 aaaggcggac ctctgccttt cgcctgggac attctgagcc ctcagtttat gtacggctcc
     3721 aaggcctacg tgaagcaccc cgccgatatt cccgactacc tgaagctgag cttccccgag
     3781 ggcttcaagt gggagagagt gatgaatttc gaggacggcg gcgtggtcac cgtgacacaa
     3841 gatagttctc tgcaggacgg cgagttcatc tacaaagtga agctgcgggg caccaacttt
     3901 ccctctgacg gacccgtgat gcagaaaaag accatgggct gggaagccag cagcgagaga
     3961 atgtaccctg aagatggcgc cctgaaaggg gagatcaagc agcggctgaa actgaaggat
     4021 ggcggccact acgacgccga agtcaagacc acatacaagg ccaagaaacc agtgcagctg
     4081 ccaggcgcct acaatgtgaa catcaagctg gacatcacca gccacaatga ggactacaca
     4141 atcgtggaac agtacgagag agccgagggg agacactcta ccggcggaat ggatgagctg
     4201 tacaagtgat aagcggccgc tcgagtctag agggcccgtt taaacccgct gatcagcctc
     4261 gactgtgcct tctagttgcc agccatctgt tgtttgcccc tcccccgtgc cttccttgac
     4321 cctggaaggt gccactccca ctgtcctttc ctaataaaat gaggaaattg catcgcattg
     4381 tctgagtagg tgtcattcta ttctgggggg tggggtgggg caggacagca agggggagga
     4441 ttgggaagac aatagcaggc atgctgggga tgcggtgggc tctatggctt ctgaggcgga
     4501 aagaaccagc tggggctcta gggggtatcc ccacgcgccc tgtagcggcg cattaagcgc
     4561 ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc
     4621 tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct
     4681 aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa
     4741 acttgattag ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg tttttcgccc
     4801 tttgacgttg gagtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact
     4861 caaccctatc tcggtctatt cttttgattt ataagggatt ttgccgattt cggcctattg
     4921 gttaaaaaat gagctgattt aacaaaaatt taacgcgaat taattctgtg gaatgtgtgt
     4981 cagttagggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat
     5041 ctcaattagt cagcaaccag gtgtggaaag tccccaggct ccccagcagg cagaagtatg
     5101 caaagcatgc atctcaatta gtcagcaacc atagtcccgc ccctaactcc gcccatcccg
     5161 cccctaactc cgcccagttc cgcccattct ccgccccatg gctgactaat tttttttatt
     5221 tatgcagagg ccgaggccgc ctctgcctct gagctattcc agaagtagtg aggaggcttt
     5281 tttggaggcc taggcttttg caaaaagctc ccgggagctt gtatatccat tttcggatct
     5341 gatcagcacg tgttgacaat taatcatcgg catagtatat cggcatagta taatacgaca
     5401 aggtgaggaa ctaaaccatg gccaagttga ccagtgccgt tccggtgctc accgcgcgcg
     5461 acgtcgccgg agcggtcgag ttctggaccg accggctcgg gttctcccgg gacttcgtgg
     5521 aggacgactt cgccggtgtg gtccgggacg acgtgaccct gttcatcagc gcggtccagg
     5581 accaggtggt gccggacaac accctggcct gggtgtgggt gcgcggcctg gacgagctgt
     5641 acgccgagtg gtcggaggtc gtgtccacga acttccggga cgcctccggg ccggccatga
     5701 ccgagatcgg cgagcagccg tgggggcggg agttcgccct gcgcgacccg gccggcaact
     5761 gcgtgcactt cgtggccgag gagcaggact gacacgtgct acgagatttc gattccaccg
     5821 ccgccttcta tgaaaggttg ggcttcggaa tcgttttccg ggacgccggc tggatgatcc
     5881 tccagcgcgg ggatctcatg ctggagttct tcgcccaccc caacttgttt attgcagctt
     5941 ataatggtta caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac
     6001 tgcattctag ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc tgtataccgt
     6061 cgacctctag ctagagcttg gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt
     6121 atccgctcac aattccacac aacatacgag ccggaagcat aaagtgtaaa gcctggggtg
     6181 cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct ttccagtcgg
     6241 gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc
     6301 gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc
     6361 ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata
     6421 acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg
     6481 cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct
     6541 caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa
     6601 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc
     6661 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt
     6721 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg
     6781 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg
     6841 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct
     6901 tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc
     6961 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg
     7021 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc
     7081 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt
     7141 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa
     7201 aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat
     7261 gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct
     7321 gactccccgt cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg
     7381 caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag
     7441 ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta
     7501 attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg
     7561 ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg
     7621 gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct
     7681 ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta
     7741 tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg
     7801 gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc
     7861 cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg
     7921 gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga
     7981 tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg
     8041 ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat
     8101 gttgaatact catactcttc ctttttcaat attattgaag catttatcag ggttattgtc
     8161 tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca
     8221 catttccccg aaaagtgcca cctgacgtc
//
