LOCUS       sequence-418123-        7481 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of heterobifunctional SasG linkers (3G).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7481)
  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 7481)
  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(3G)-mCherry
FEATURES             Location/Qualifiers
     source          1..7481
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(57..76)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        248..627
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        628..831
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     782..802
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     876..895
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        876..894
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             join(977..979,980..1690)
                     /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(3502..3519)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3508..3732
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3778..4206
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3865..3884)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4075..4096
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4215..4235)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4220..4549
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4400..4535
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4462..4481
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        4597..4644
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             4663..5037
                     /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    5167..5288
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5204..5223)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5258..5277
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5337..5353)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5337..5353)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5350..5372)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5361..5377
                     /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(5385..5391,5392..5409,5410..5415))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5430..5451
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5568..5585)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5739..6327)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5819..5838)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6498..7289,7290..7358))
                     /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     7121..7140
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7359..7463)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gccacctgac gtcgacggat cgggagatct cccgatcccc tatggtgcac tctcagtaca
       61 atctgctctg atgccgcata gttaagccag tatctgctcc ctgcttgtgt gttggaggtc
      121 gctgagtagt gcgcgagcaa aatttaagct acaacaaggc aaggcttgac cgacaattgc
      181 atgaagaatc tgcttagggt taggcgtttt gcgctgcttc gcgatgtacg ggccagatat
      241 acgcgttgac attgattatt gactagttat taatagtaat caattacggg gtcattagtt
      301 catagcccat atatggagtt ccgcgttaca taacttacgg taaatggccc gcctggctga
      361 ccgcccaacg acccccgccc attgacgtca ataatgacgt atgttcccat agtaacgcca
      421 atagggactt tccattgacg tcaatgggtg gagtatttac ggtaaactgc ccacttggca
      481 gtacatcaag tgtatcatat gccaagtacg ccccctattg acgtcaatga cggtaaatgg
      541 cccgcctggc attatgccca gtacatgacc ttatgggact ttcctacttg gcagtacatc
      601 tacgtattag tcatcgctat taccatggtg atgcggtttt ggcagtacat caatgggcgt
      661 ggatagcggt ttgactcacg gggatttcca agtctccacc ccattgacgt caatgggagt
      721 ttgttttggc accaaaatca acgggacttt ccaaaatgtc gtaacaactc cgccccattg
      781 acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata taagcagagc tctctggcta
      841 actagagaac ccactgctta ctggcttatc gaaattaata cgactcacta tagggagacc
      901 caagctggct agcgtttaaa cttaagcttg gtaccgagct cggatccact agtccagtgt
      961 ggtggaattc gccaccatga gcaagggcga agaactgttt accggcgtgg tgcccatcct
     1021 ggtggaactg gatggggatg tgaacggcca caagttcagc gtcagaggcg aaggcgaagg
     1081 ggatgccaca aacggcaagc tgaccctgaa gttcatctgc accaccggaa agctgcccgt
     1141 gccttggcct acactggtca ccacactgac atacggcgtg cagtgcttca gcagataccc
     1201 cgaccatatg aagcggcacg acttcttcaa gagcgccatg cctgagggct acgtgcaaga
     1261 gagaaccatc agcttcaagg acgacggcac ctacaagacc agagccgaag tgaagttcga
     1321 gggcgacacc ctggtcaacc ggatcgagct gaagggcatc gatttcaaag aggacggcaa
     1381 catcctgggc cacaagcttg agtacaactt caacagccac aacgtgtaca tcaccgccga
     1441 caagcagaag aacggcatca aggccaactt caagatccgg cacaacgtgg aagatggcag
     1501 cgtgcagctg gccgatcact accagcagaa cacacccatc ggagatggcc ctgtgctgct
     1561 gcccgataac cactacctga gcacacagag cgtgctgagc aaggacccca acgagaagag
     1621 ggaccacatg gtgctgctgg agtttgtgac agccgccgga atcacccacg gcatggacga
     1681 actttacaaa ggcggcaccg gtggcagcgc ccctaagaca attaccgagc tggaaaagaa
     1741 ggtggaagag atccccttca agaaagagcg gaagttcaac cccgatctgg cccctggcac
     1801 cgagaaagtg acaagagaag gccagaaggg cgagaaaacc atcaccacac ctacactgaa
     1861 gaaccctctg accggcgtga tcatctccaa gggcgagccc aaagaggaaa tcacaaagga
     1921 ccctatcaac gagctgaccg agtacggccc cgagacaatt gctcctggcc acagagatga
     1981 gttcgacccc aaactgccca ccggcgagaa agaagaagtg cctggcaagc ctggcatcaa
     2041 gaaccctgag acaggcgacg tcgtcagacc tcctgtggac agcgtgacca aatacggccc
     2101 tgtgaagggc gacagcatcg tggaaaaaga agagattccg tttgagaaag agagaaagtt
     2161 taatcccgac ctcgctccag ggacagaaaa agtcactcgc gagggccaga aaggggaaaa
     2221 gacaatcaca accccaacgc tgaaaaatcc cctgaccggg gagattatca gcaaaggcga
     2281 gagcaaagaa gaaattacga aagatcccat caatgaactc acagagtacg ggcccgagac
     2341 aattgctcct ggccacagag atgagttcga ccccaaactg cccaccggcg agaaagaaga
     2401 agtgcctggc aagcctggca tcaagaaccc tgagacaggc gacgtcgtca gacctcctgt
     2461 ggacagcgtg accaaatacg gccctgtgaa gggcgacagc atcgtggaaa aagaagagat
     2521 tccgtttgag aaagagagaa agtttaatcc cgacctcgct ccagggacag aaaaagtcac
     2581 tcgcgagggc cagaaagggg aaaagacaat cacaacccca acgctgaaaa atcccctgac
     2641 cggggagatt atcagcaaag gcgagagcaa agaagaaatt acgaaagatc ccatcaatga
     2701 actcacagag tacgggcctg agacgggctc cggagccaca ggatctgtgt ccaaaggcga
     2761 agaggacaac atggccatca tcaaagaatt tatgcggttc aaggtgcaca tggaaggctc
     2821 cgtgaatggc cacgagtttg agattgaagg cgaaggcgag ggcagaccct atgagggaac
     2881 acagaccgcc aagctgaaag tgaccaaagg cggacctctg cctttcgcct gggacattct
     2941 gagccctcag tttatgtacg gctccaaggc ctacgtgaag caccccgccg atattcccga
     3001 ctacctgaag ctgagcttcc ccgagggctt caagtgggag agagtgatga atttcgagga
     3061 cggcggcgtg gtcaccgtga cacaagatag ttctctgcag gacggcgagt tcatctacaa
     3121 agtgaagctg cggggcacca actttccctc tgacggaccc gtgatgcaga aaaagaccat
     3181 gggctgggaa gccagcagcg agagaatgta ccctgaagat ggcgccctga aaggggagat
     3241 caagcagcgg ctgaaactga aggatggcgg ccactacgac gccgaagtca agaccacata
     3301 caaggccaag aaaccagtgc agctgccagg cgcctacaat gtgaacatca agctggacat
     3361 caccagccac aatgaggact acacaatcgt ggaacagtac gagagagccg aggggagaca
     3421 ctctaccggc ggaatggatg agctgtacaa gtgataagcg gccgctcgag tctagagggc
     3481 ccgtttaaac ccgctgatca gcctcgactg tgccttctag ttgccagcca tctgttgttt
     3541 gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc ctttcctaat
     3601 aaaatgagga aattgcatcg cattgtctga gtaggtgtca ttctattctg gggggtgggg
     3661 tggggcagga cagcaagggg gaggattggg aagacaatag caggcatgct ggggatgcgg
     3721 tgggctctat ggcttctgag gcggaaagaa ccagctgggg ctctaggggg tatccccacg
     3781 cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta
     3841 cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt
     3901 tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg
     3961 ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt agtgggccat
     4021 cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac
     4081 tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt gatttataag
     4141 ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg
     4201 cgaattaatt ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag gctccccagc
     4261 aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc
     4321 aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccatagt
     4381 cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc attctccgcc
     4441 ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctctg cctctgagct
     4501 attccagaag tagtgaggag gcttttttgg aggcctaggc ttttgcaaaa agctcccggg
     4561 agcttgtata tccattttcg gatctgatca gcacgtgttg acaattaatc atcggcatag
     4621 tatatcggca tagtataata cgacaaggtg aggaactaaa ccatggccaa gttgaccagt
     4681 gccgttccgg tgctcaccgc gcgcgacgtc gccggagcgg tcgagttctg gaccgaccgg
     4741 ctcgggttct cccgggactt cgtggaggac gacttcgccg gtgtggtccg ggacgacgtg
     4801 accctgttca tcagcgcggt ccaggaccag gtggtgccgg acaacaccct ggcctgggtg
     4861 tgggtgcgcg gcctggacga gctgtacgcc gagtggtcgg aggtcgtgtc cacgaacttc
     4921 cgggacgcct ccgggccggc catgaccgag atcggcgagc agccgtgggg gcgggagttc
     4981 gccctgcgcg acccggccgg caactgcgtg cacttcgtgg ccgaggagca ggactgacac
     5041 gtgctacgag atttcgattc caccgccgcc ttctatgaaa ggttgggctt cggaatcgtt
     5101 ttccgggacg ccggctggat gatcctccag cgcggggatc tcatgctgga gttcttcgcc
     5161 caccccaact tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat
     5221 ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat
     5281 gtatcttatc atgtctgtat accgtcgacc tctagctaga gcttggcgta atcatggtca
     5341 tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat acgagccgga
     5401 agcataaagt gtaaagcctg gggtgcctaa tgagtgagct aactcacatt aattgcgttg
     5461 cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc
     5521 caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc gctcactgac
     5581 tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata
     5641 cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa
     5701 aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct
     5761 gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa
     5821 agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg
     5881 cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca
     5941 cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa
     6001 ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg
     6061 gtaagacacg acttatcgcc actggcagca gccactggta acaggattag cagagcgagg
     6121 tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga
     6181 acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc
     6241 tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag
     6301 attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac
     6361 gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc
     6421 ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag
     6481 taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt
     6541 ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag
     6601 ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca
     6661 gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact
     6721 ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca
     6781 gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg
     6841 tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc
     6901 atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg
     6961 gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca
     7021 tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt
     7081 atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc
     7141 agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc
     7201 ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca
     7261 tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa
     7321 aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat
     7381 tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa
     7441 aataaacaaa taggggttcc gcgcacattt ccccgaaaag t
//
