LOCUS       sequence-440205-        5950 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, 20 GS linker, CyOFP1 protein..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5950)
  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 5950)
  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: L2127-FRB-GPA-114-20-CyOFP1
FEATURES             Location/Qualifiers
     source          1..5950
                     /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(2573..2590)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2579..2803
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2849..3277
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2936..2955)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3146..3167
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        3300..3496
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      3347..3482
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3409..3428
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     polyA_signal    3616..3737
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3653..3672)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3707..3726
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3786..3802)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3786..3802)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3799..3821)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3810..3826
                     /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(3834..3840,3841..3858,3859..3864))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3879..3900
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4017..4034)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4188..4776)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4268..4287)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4947..5738,5739..5807))
                     /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     5570..5589
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5808..5912)
                     /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 cggggggtct ggaggcagtg
     1801 gtgggtctgg tggtggttct ggggggagtg ggggttcagg cgagctgatc aaagaaaaca
     1861 tgcggagcaa gctgtacctg gaaggcagcg tgaacggcca ccagttcaag tgtacacacg
     1921 aaggcgaggg caagccctac gaaggcaagc agaccaaccg gattaaggtg gtggaaggcg
     1981 gccctctgcc tttcgccttt gatatcctgg ccacacactt tatgtacggc agcaaggtgt
     2041 tcatcaagta ccccgccgac ctgcctgact acttcaagca gagcttcccc gagggcttca
     2101 cctgggagcg agtgatggtg tttgaggacg gcggagtgct gaccgccaca caggatactt
     2161 ctctgcagga cggcgagctg atctacaacg tgaaagtgcg gggcgtgaac ttccccgcca
     2221 atggacctgt gatgcagaag aaaaccctcg gctgggagcc cagcaccgag acaatgtatc
     2281 cagccgatgg cggcctcgag ggcagatgtg ataaggccct gaaactcgtc ggcggaggcc
     2341 atctgcacgt gaactttaag accacctaca agagcaagaa acccgtgaag atgcccggcg
     2401 tgcactacgt ggacagacgg ctggaaagaa tcaaagaggc cgacaacgag acatacgtgg
     2461 aacagtacga gcacgccgtg gccagataca gcaatctcgg aggcggaatg gacgagctgt
     2521 acaaatgagc ggccgctcga gtctagaggg cccgtttaaa cccgctgatc agcctcgact
     2581 gtgccttcta gttgccagcc atctgttgtt tgcccctccc ccgtgccttc cttgaccctg
     2641 gaaggtgcca ctcccactgt cctttcctaa taaaatgagg aaattgcatc gcattgtctg
     2701 agtaggtgtc attctattct ggggggtggg gtggggcagg acagcaaggg ggaggattgg
     2761 gaagtcaata gcaggcatgc tggggatgcg gtgggctcta tggcttctga ggcggaaaga
     2821 accagctggg gctctagggg gtatccccac gcgccctgta gcggcgcatt aagcgcggcg
     2881 ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca gcgccctagc gcccgctcct
     2941 ttcgctttct tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat
     3001 cgggggctcc ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt
     3061 gattagggtg atggttcacg tagtgggcca tcgccctgat agacggtttt tcgccctttg
     3121 acgttggagt ccacgttctt taatagtgga ctcttgttcc aaactggaac aacactcaac
     3181 cctatctcgg tctattcttt tgatttataa gggattttgc cgatttcggc ctattggtta
     3241 aaaaatgagc tgatttaaca aaaatttaac gcgaattaat tctgtggaat gtgtgtcagt
     3301 taggctcaat tagtcagcaa ccatagtccc gcccctaact ccgcccatcc cgcccctaac
     3361 tccgcccagt tccgcccatt ctccgcccca tggctgacta atttttttta tttatgcaga
     3421 ggccgaggcc gcctctgcct ctgagctatt ccagaagtag tgaggaggct tttttggagg
     3481 cctaggcttt tgcaaaaagc tcccgggagc ttgtatatcc attttcggag gttgggcttc
     3541 ggaatcgttt tccgggacgc cggctggatg atcctccagc gcggggatct catgctggag
     3601 ttcttcgccc accccaactt gtttattgca gcttataatg gttacaaata aagcaatagc
     3661 atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa
     3721 ctcatcaatg tatcttatca tgtctgtata ccgtcgacct ctagctagag cttggcgtaa
     3781 tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata
     3841 cgagccggaa gcataaagtg taaagcctgg ggtgcctaat gagtgagcta actcacatta
     3901 attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa
     3961 tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg
     4021 ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag
     4081 gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa
     4141 ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc
     4201 cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca
     4261 ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg
     4321 accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct
     4381 catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt
     4441 gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag
     4501 tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc
     4561 agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac
     4621 actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga
     4681 gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc
     4741 aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg
     4801 gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca
     4861 aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt
     4921 atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca
     4981 gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg
     5041 atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga tccacgctca
     5101 ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt
     5161 cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt
     5221 agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca
     5281 cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca
     5341 tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga
     5401 agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact
     5461 gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga
     5521 gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg
     5581 ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc
     5641 tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga
     5701 tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat
     5761 gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt
     5821 caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt
     5881 atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctgac
     5941 gtcgacggat
//
