LOCUS       sequence-409224-        8633 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of heterobifunctional SasG linkers (6G).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8633)
  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 8633)
  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(6G)-mCherry
FEATURES             Location/Qualifiers
     source          1..8633
                     /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(4641..4658)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4647..4871
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      4917..5345
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5004..5023)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5214..5235
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5354..5374)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        5359..5688
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5539..5674
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     5601..5620
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        5736..5783
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             5802..6176
                     /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    6306..6427
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6343..6362)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6397..6416
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(6476..6492)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6476..6492)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6489..6511)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6500..6516
                     /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(6524..6530,6531..6548,6549..6554))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6569..6590
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6707..6724)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6878..7466)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6958..6977)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7637..8428,8429..8497))
                     /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     8260..8279
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8498..8602)
                     /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 gcccgagaca
     3481 attgctcctg gccacagaga tgagttcgac cccaaactgc ccaccggcga gaaagaagaa
     3541 gtgcctggca agcctggcat caagaaccct gagacaggcg acgtcgtcag acctcctgtg
     3601 gacagcgtga ccaaatacgg ccctgtgaag ggcgacagca tcgtggaaaa agaagagatt
     3661 ccgtttgaga aagagagaaa gtttaatccc gacctcgctc cagggacaga aaaagtcact
     3721 cgcgagggcc agaaagggga aaagacaatc acaaccccaa cgctgaaaaa tcccctgacc
     3781 ggggagatta tcagcaaagg cgagagcaaa gaagaaatta cgaaagatcc catcaatgaa
     3841 ctcacagagt acgggcctga gacgggctcc ggagccacag gatctgtgtc caaaggcgaa
     3901 gaggacaaca tggccatcat caaagaattt atgcggttca aggtgcacat ggaaggctcc
     3961 gtgaatggcc acgagtttga gattgaaggc gaaggcgagg gcagacccta tgagggaaca
     4021 cagaccgcca agctgaaagt gaccaaaggc ggacctctgc ctttcgcctg ggacattctg
     4081 agccctcagt ttatgtacgg ctccaaggcc tacgtgaagc accccgccga tattcccgac
     4141 tacctgaagc tgagcttccc cgagggcttc aagtgggaga gagtgatgaa tttcgaggac
     4201 ggcggcgtgg tcaccgtgac acaagatagt tctctgcagg acggcgagtt catctacaaa
     4261 gtgaagctgc ggggcaccaa ctttccctct gacggacccg tgatgcagaa aaagaccatg
     4321 ggctgggaag ccagcagcga gagaatgtac cctgaagatg gcgccctgaa aggggagatc
     4381 aagcagcggc tgaaactgaa ggatggcggc cactacgacg ccgaagtcaa gaccacatac
     4441 aaggccaaga aaccagtgca gctgccaggc gcctacaatg tgaacatcaa gctggacatc
     4501 accagccaca atgaggacta cacaatcgtg gaacagtacg agagagccga ggggagacac
     4561 tctaccggcg gaatggatga gctgtacaag tgataagcgg ccgctcgagt ctagagggcc
     4621 cgtttaaacc cgctgatcag cctcgactgt gccttctagt tgccagccat ctgttgtttg
     4681 cccctccccc gtgccttcct tgaccctgga aggtgccact cccactgtcc tttcctaata
     4741 aaatgaggaa attgcatcgc attgtctgag taggtgtcat tctattctgg ggggtggggt
     4801 ggggcaggac agcaaggggg aggattggga agacaatagc aggcatgctg gggatgcggt
     4861 gggctctatg gcttctgagg cggaaagaac cagctggggc tctagggggt atccccacgc
     4921 gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac
     4981 acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt
     5041 cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc
     5101 tttacggcac ctcgacccca aaaaacttga ttagggtgat ggttcacgta gtgggccatc
     5161 gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact
     5221 cttgttccaa actggaacaa cactcaaccc tatctcggtc tattcttttg atttataagg
     5281 gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc
     5341 gaattaattc tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg ctccccagca
     5401 ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca
     5461 ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc aaccatagtc
     5521 ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca ttctccgccc
     5581 catggctgac taattttttt tatttatgca gaggccgagg ccgcctctgc ctctgagcta
     5641 ttccagaagt agtgaggagg cttttttgga ggcctaggct tttgcaaaaa gctcccggga
     5701 gcttgtatat ccattttcgg atctgatcag cacgtgttga caattaatca tcggcatagt
     5761 atatcggcat agtataatac gacaaggtga ggaactaaac catggccaag ttgaccagtg
     5821 ccgttccggt gctcaccgcg cgcgacgtcg ccggagcggt cgagttctgg accgaccggc
     5881 tcgggttctc ccgggacttc gtggaggacg acttcgccgg tgtggtccgg gacgacgtga
     5941 ccctgttcat cagcgcggtc caggaccagg tggtgccgga caacaccctg gcctgggtgt
     6001 gggtgcgcgg cctggacgag ctgtacgccg agtggtcgga ggtcgtgtcc acgaacttcc
     6061 gggacgcctc cgggccggcc atgaccgaga tcggcgagca gccgtggggg cgggagttcg
     6121 ccctgcgcga cccggccggc aactgcgtgc acttcgtggc cgaggagcag gactgacacg
     6181 tgctacgaga tttcgattcc accgccgcct tctatgaaag gttgggcttc ggaatcgttt
     6241 tccgggacgc cggctggatg atcctccagc gcggggatct catgctggag ttcttcgccc
     6301 accccaactt gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt
     6361 tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg
     6421 tatcttatca tgtctgtata ccgtcgacct ctagctagag cttggcgtaa tcatggtcat
     6481 agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa
     6541 gcataaagtg taaagcctgg ggtgcctaat gagtgagcta actcacatta attgcgttgc
     6601 gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc
     6661 aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact
     6721 cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac
     6781 ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa
     6841 aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg
     6901 acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa
     6961 gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc
     7021 ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac
     7081 gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac
     7141 cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg
     7201 taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt
     7261 atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa
     7321 cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct
     7381 cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga
     7441 ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg
     7501 ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct
     7561 tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt
     7621 aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc
     7681 tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg
     7741 gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag
     7801 atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt
     7861 tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag
     7921 ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt
     7981 ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca
     8041 tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg
     8101 ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat
     8161 ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta
     8221 tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca
     8281 gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct
     8341 taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat
     8401 cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa
     8461 agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt
     8521 gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa
     8581 ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctgac gtc
//
