LOCUS       sequence-440200-        5920 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Biosensor chain detecting rapamycin and responding with BRET. FRB 
            ectodomain, human Glycophorin A (GPA) scaffold, 20 GS linker, split 
            Nanoluciferase fragment 114, 10 GS linker, CyOFP1 protein..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5920)
  AUTHORS   Dolberg TB, Gunnels TF, Ling T, Sarnese KA, Crispino JD, Leonard JN
  TITLE     Building Synthetic Biosensors Using Red Blood Cell Proteins.
  JOURNAL   ACS Synth Biol. 2024 Apr 19;13(4):1273-1289. doi: 
            10.1021/acssynbio.3c00754. Epub 2024 Mar 27.
  PUBMED    38536408
REFERENCE   2  (bases 1 to 5920)
  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"; doi: 
            "10.1021/acssynbio.3c00754"; journalName: "ACS Synth Biol"; date: 
            "2024-04-19- 19"; volume: "13"; issue: "4"; pages: "1273-1289"
COMMENT     Sequence Label: L2128-FRB-GPA-114-10-CyOFP1
FEATURES             Location/Qualifiers
     source          1..5920
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(37..56)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        228..607
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        608..811
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     762..782
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     856..875
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        856..874
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             957..986
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     primer_bind     complement(2543..2560)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2549..2773
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2819..3247
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2906..2925)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3116..3137
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        3270..3466
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      3317..3452
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3379..3398
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     polyA_signal    3586..3707
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3623..3642)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3677..3696
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3756..3772)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3756..3772)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3769..3791)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3780..3796
                     /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(3804..3810,3811..3828,3829..3834))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3849..3870
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3987..4004)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4158..4746)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4238..4257)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4917..5708,5709..5777))
                     /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     5540..5559
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5778..5882)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 cgggagatct cccgatcccc tatggtgcac tctcagtaca atctgctctg atgccgcata
       61 gttaagccag tatctgctcc ctgcttgtgt gttggaggtc gctgagtagt gcgcgagcaa
      121 aatttaagct acaacaaggc aaggcttgac cgacaattgc atgaagaatc tgcttagggt
      181 taggcgtttt gcgctgcttc gcgatgtacg ggccagatat acgcgttgac attgattatt
      241 gactagttat taatagtaat caattacggg gtcattagtt catagcccat atatggagtt
      301 ccgcgttaca taacttacgg taaatggccc gcctggctga ccgcccaacg acccccgccc
      361 attgacgtca ataatgacgt atgttcccat agtaacgcca atagggactt tccattgacg
      421 tcaatgggtg gagtatttac ggtaaactgc ccacttggca gtacatcaag tgtatcatat
      481 gccaagtacg ccccctattg acgtcaatga cggtaaatgg cccgcctggc attatgccca
      541 gtacatgacc ttatgggact ttcctacttg gcagtacatc tacgtattag tcatcgctat
      601 taccatggtg atgcggtttt ggcagtacat caatgggcgt ggatagcggt ttgactcacg
      661 gggatttcca agtctccacc ccattgacgt caatgggagt ttgttttggc accaaaatca
      721 acgggacttt ccaaaatgtc gtaacaactc cgccccattg acgcaaatgg gcggtaggcg
      781 tgtacggtgg gaggtctata taagcagagc tctctggcta actagagaac ccactgctta
      841 ctggcttatc gaaattaata cgactcacta tagggagtcc caagctggct agcgccacca
      901 tgtatggaaa aataatcttt gtattactat tgtcagaaat tgtgagcata tcagcagagc
      961 agaaactcat ctcagaagag gatctgggcg cgcctatgct cgagatgtgg catgaaggcc
     1021 tggaagaggc atctcgtttg tactttgggg aaaggaacgt gaaaggcatg tttgaggtgc
     1081 tggagccctt gcatgctatg atggaacggg gcccccagac tctgaaggaa acatccttta
     1141 atcaggccta tggtcgagat ttaatggagg cccaagagtg gtgcaggaag tacatgaaat
     1201 cagggaatgt caaggacctc ctccaagcct gggacctcta ttatcatgtg ttccgacgaa
     1261 tctcaaagac tagaaccggt ttaagtacca ctgaggtggc aatgcacact tcaacctctt
     1321 cttcagtcac aaagagttac atctcatcac agacaaatga tacgcacaaa cgggacacat
     1381 atgcagccac tcctagagct catgaagttt cagaaatttc tgttagaact gtttaccctc
     1441 cagaagagga aaccggagaa agggtacaac ttgcccatca tttctctgaa ccagagataa
     1501 cactcattat ttttggggtg atggctggtg ttattggaac gatcctctta atttcttacg
     1561 gtattcgccg actgataaag aaaagcccat ctgatgtaaa acctctcccc tcacctgaca
     1621 cagacgtgcc tttaagttct gttgaaatag aaaatccaga gacaagtgat caattaatta
     1681 agggcggcag cggcggctcc ggtggatctg gcggtggaag cggaggttct ggtggttctg
     1741 gcgtgacagg ctaccggctg ttcgaggaaa tcctgggcgc cggtggttct ggggggagtg
     1801 ggggttcagg cgagctgatc aaagaaaaca tgcggagcaa gctgtacctg gaaggcagcg
     1861 tgaacggcca ccagttcaag tgtacacacg aaggcgaggg caagccctac gaaggcaagc
     1921 agaccaaccg gattaaggtg gtggaaggcg gccctctgcc tttcgccttt gatatcctgg
     1981 ccacacactt tatgtacggc agcaaggtgt tcatcaagta ccccgccgac ctgcctgact
     2041 acttcaagca gagcttcccc gagggcttca cctgggagcg agtgatggtg tttgaggacg
     2101 gcggagtgct gaccgccaca caggatactt ctctgcagga cggcgagctg atctacaacg
     2161 tgaaagtgcg gggcgtgaac ttccccgcca atggacctgt gatgcagaag aaaaccctcg
     2221 gctgggagcc cagcaccgag acaatgtatc cagccgatgg cggcctcgag ggcagatgtg
     2281 ataaggccct gaaactcgtc ggcggaggcc atctgcacgt gaactttaag accacctaca
     2341 agagcaagaa acccgtgaag atgcccggcg tgcactacgt ggacagacgg ctggaaagaa
     2401 tcaaagaggc cgacaacgag acatacgtgg aacagtacga gcacgccgtg gccagataca
     2461 gcaatctcgg aggcggaatg gacgagctgt acaaatgagc ggccgctcga gtctagaggg
     2521 cccgtttaaa cccgctgatc agcctcgact gtgccttcta gttgccagcc atctgttgtt
     2581 tgcccctccc ccgtgccttc cttgaccctg gaaggtgcca ctcccactgt cctttcctaa
     2641 taaaatgagg aaattgcatc gcattgtctg agtaggtgtc attctattct ggggggtggg
     2701 gtggggcagg acagcaaggg ggaggattgg gaagtcaata gcaggcatgc tggggatgcg
     2761 gtgggctcta tggcttctga ggcggaaaga accagctggg gctctagggg gtatccccac
     2821 gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct
     2881 acacttgcca gcgccctagc gcccgctcct ttcgctttct tcccttcctt tctcgccacg
     2941 ttcgccggct ttccccgtca agctctaaat cgggggctcc ctttagggtt ccgatttagt
     3001 gctttacggc acctcgaccc caaaaaactt gattagggtg atggttcacg tagtgggcca
     3061 tcgccctgat agacggtttt tcgccctttg acgttggagt ccacgttctt taatagtgga
     3121 ctcttgttcc aaactggaac aacactcaac cctatctcgg tctattcttt tgatttataa
     3181 gggattttgc cgatttcggc ctattggtta aaaaatgagc tgatttaaca aaaatttaac
     3241 gcgaattaat tctgtggaat gtgtgtcagt taggctcaat tagtcagcaa ccatagtccc
     3301 gcccctaact ccgcccatcc cgcccctaac tccgcccagt tccgcccatt ctccgcccca
     3361 tggctgacta atttttttta tttatgcaga ggccgaggcc gcctctgcct ctgagctatt
     3421 ccagaagtag tgaggaggct tttttggagg cctaggcttt tgcaaaaagc tcccgggagc
     3481 ttgtatatcc attttcggag gttgggcttc ggaatcgttt tccgggacgc cggctggatg
     3541 atcctccagc gcggggatct catgctggag ttcttcgccc accccaactt gtttattgca
     3601 gcttataatg gttacaaata aagcaatagc atcacaaatt tcacaaataa agcatttttt
     3661 tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca tgtctgtata
     3721 ccgtcgacct ctagctagag cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat
     3781 tgttatccgc tcacaattcc acacaacata cgagccggaa gcataaagtg taaagcctgg
     3841 ggtgcctaat gagtgagcta actcacatta attgcgttgc gctcactgcc cgctttccag
     3901 tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt
     3961 ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg
     4021 ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg
     4081 gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag
     4141 gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga
     4201 cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct
     4261 ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc
     4321 tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg
     4381 gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc
     4441 tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca
     4501 ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag
     4561 ttcttgaagt ggtggcctaa ctacggctac actagaagaa cagtatttgg tatctgcgct
     4621 ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc
     4681 accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga
     4741 tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca
     4801 cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat
     4861 taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac
     4921 caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt
     4981 gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt
     5041 gctgcaatga taccgcgaga tccacgctca ccggctccag atttatcagc aataaaccag
     5101 ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct
     5161 attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt
     5221 gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc
     5281 tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt
     5341 agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg
     5401 gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg
     5461 actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct
     5521 tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc
     5581 attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt
     5641 tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt
     5701 tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg
     5761 aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta tcagggttat
     5821 tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg
     5881 cgcacatttc cccgaaaagt gccacctgac gtcgacggat
//
