LOCUS       sequence-436396-        5230 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Caging at cytosolic C-ter of POI: mScarletCD4-PhoCl-RER in 
            pTREtight2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5230)
  AUTHORS   Kashyap P, Bertelli S, Cao F, Kostritskaia Y, Blank F, Srikanth NA, 
            Schlack-Leigers C, Saleppico R, Bierhuizen D, Lu X, Nickel W, 
            Campbell RE, Plested AJR, Stauber T, Taylor MJ, Ewers H
  TITLE     An optogenetic method for the controlled release of single 
            molecules.
  JOURNAL   Nat Methods. 2024 Mar 8. doi: 10.1038/s41592-024-02204-x.
  PUBMED    38459384
REFERENCE   2  (bases 1 to 5230)
  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 
            Methods. 2024 Mar 8. doi: 10.1038/s41592-024-02204-x."
COMMENT     Sequence Label: pInsert-PhoCl-RER
FEATURES             Location/Qualifiers
     source          1..5230
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             821..856
                     /codon_start=1
                     /product="nanobody epitope tag corresponding to amino acids
                     16-27 of human beta-catenin (Traenkle et al., 2015)"
                     /label=BC2 tag
                     /translation="PDRKAAVSHWQQ"
     polyA_signal    2502..2583
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2539..2558)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2686..2708
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2746..2764)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2832..2936
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2937..3005,3006..3797)
                     /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     complement(3155..3174)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3968..4556
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4457..4476
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4710..4727
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    4844..4865
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(4880..4885,4886..4903,4904..4910)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     promoter        join(4919..5230,1..3)
                     /label=tight TRE promoter
                     /note="Tet-responsive promoter PTight, consisting of seven 
                     tet operator sequences followed by the minimal CMV 
                     promoter"
     protein_bind    4927..4945
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    4963..4981
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    4998..5016
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    5034..5052
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    5070..5088
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    5105..5123
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    5141..5159
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     5206..5230
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
ORIGIN
        1 gcctggagaa ttcgagctcg gtacccgggg atcctctagt cagctgacgc gtgctagcgg
       61 atccatgtgg cctcttgttg ctgctctgct gctgggctct gcctgttgtg gatctgccgg
      121 ctccatggtg tctaagggcg aagccgtgat caaagaattc atgcggttca aggtgcacat
      181 ggaaggcagc atgaacggcc acgagttcga gatcgaaggc gaaggcgagg gcagacctta
      241 tgagggaaca cagaccgcca agctgaaagt gaccaaaggc ggccctctgc ctttcagctg
      301 ggacattctg agccctcagt ttatgtacgg cagccgggcc ttcacaaagc accctgccga
      361 tattcccgac tactacaagc agagcttccc cgagggcttc aagtgggaga gagtgatgaa
      421 cttcgaggac ggcggagccg tgaccgtgac acaggataca agcctggaag atggcaccct
      481 gatctacaaa gtgaagctgc ggggcaccaa ctttccacct gatggccccg tgatgcagaa
      541 aaagaccatg ggctgggaag ccagcaccga gagactgtat cctgaggatg gcgtgctgaa
      601 gggcgacatc aagatggccc tgagactgaa ggatggcggc agatacctgg ccgacttcaa
      661 gaccacctac aaggccaaga aacccgtgca gatgcctggc gcctacaacg tggacagaaa
      721 gctggacatc accagccaca acgaggacta caccgtggtg gaacagtacg agcggagcga
      781 aggcagacac tctacaggcg gaatggacga gctgtacaag cccgatagaa aggccgccgt
      841 gtctcactgg cagcaaggat ctggcctcga gagcacacct gtgcagccta tggctctgat
      901 tgtgcttggc ggagtggctg gcctgctgct gtttatcggc ctgggcatct tcttttgcgt
      961 gcggtgcaga cacagaagaa gaggcagcgg caagcttgtc atacccgact acttcaagca
     1021 gagcttcccc gagggctaca gctgggagag atccatgaca tacgaggacg gcggcatctg
     1081 tatcgccacc aacgacatca ccatggaagg cgacagcttc atcaacaaga tccactttca
     1141 gggcacgaac ttccctccaa acggccccgt gatgcagaag agaacagttg gctgggaagc
     1201 cagcaccgag aagatgtacg agcgagatgg cgtgctgaag ggcgacgtga aaatgaagct
     1261 gctgctgaaa ggcggcggac actacagagg cgactaccgg accacctaca aagtgaagca
     1321 gaaacccgtg aagctgcccg actgccactt cgtggatcac cggatcgaga tcctgagcca
     1381 cgacaaggac tacaacaaag tcaagctgta cgagcacgcc gtggccaaga cctccaccga
     1441 ttctatggac gagctgtaca aaggcggcag cggcggcatg gtgtctaagg gcgaagagac
     1501 aatcaccagc gtgatcaagc ccgacatgaa gaacaagctg cggatggaag gcaacgtgaa
     1561 cggccacgcc tttgtgatcg aaggcgaagg cagcggcaag ccctttgagg gcatccagac
     1621 catcgacctg gaagtgaaag aaggcgcccc tctgcctttc gcctacgaca ttctgaccac
     1681 cgccttccac tacggcaacc gggtgttcac aaagtaccct agaggcggag gcggcacagg
     1741 atctggcccg cggatgtctg aaggcgattc tgtgggagag tctgtgcacg gcaagcctag
     1801 cgtggtgtac cggttcttta cccggctggg ccagatctac cagagctggc tggataagag
     1861 caccccttac accgccgtca gatgggttgt gacactgggc ctgagcttcg tgtacatgat
     1921 ccgggtgtac ctgctgcaag gctggtacat cgtgacatac gccctgggaa tctaccacct
     1981 gaacctgttc attgcctttc tgtctcccaa ggtggacccc agcctgatgg aagattccga
     2041 cgatggacct agcctgccta ccaagcagaa cgaggaattc agacccttca tccggcggct
     2101 gcccgagttc aaattttggc acgccgccac caagggcatc ctggttgcta tggtctgcac
     2161 cttcttcgac gccttcaacg tgcccgtgtt ctggcccatc ctggtcatgt acttcatcat
     2221 gctgttctgt atcaccatga agcgccagat caagcacatg atcaagtacc gctacatccc
     2281 cttcacacac ggcaagaggc ggtacagagg caaagaggat gccggaaaag ccttcgccag
     2341 ctaatctaga gctagcgcgg ccgcatcgat aagcttgtcg acgatatctc tagaggatca
     2401 taatcagcca taccacattt gtagaggttt tacttgcttt aaaaaacctc ccacacctcc
     2461 ccctgaacct gaaacataaa atgaatgcaa ttgttgttgt taacttgttt attgcagctt
     2521 ataatggtta caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac
     2581 tgcctcgaga gttaagccag ccccgacacc cgccaacacc cgctgacgcg ccctgacggg
     2641 cttgtctgct cccggcatcc gcttacagac aagctgtgac cgtctccggg agctgcatgt
     2701 gtcagaggtt ttcaccgtca tcaccgaaac gcgcgagacg aaagggcctc gtgatacgcc
     2761 tatttttata ggttaatgtc atgataataa tggtttctta gacgtcaggt ggcacttttc
     2821 ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc
     2881 cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtatga
     2941 gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt
     3001 ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcacgag
     3061 tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag
     3121 aacgttttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta ttatcccgta
     3181 ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg
     3241 agtactcacc agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca
     3301 gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca acgatcggag
     3361 gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc
     3421 gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg
     3481 tagcaatggc aacaacgttg cgcaaactat taactggcga actacttact ctagcttccc
     3541 ggcaacaatt aatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg
     3601 cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt gggtctcgcg
     3661 gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga
     3721 cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac
     3781 tgattaagca ttggtaactg tcagaccaag tttactcata tatactttag attgatttaa
     3841 aacttcattt ttaatttaaa aggatctagg tgaagatcct ttttgataat ctcatgacca
     3901 aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag
     3961 gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac
     4021 cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa
     4081 ctggcttcag cagagcgcag ataccaaata ctgttcttct agtgtagccg tagttaggcc
     4141 accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag
     4201 tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac
     4261 cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc
     4321 gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc
     4381 ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca
     4441 cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc
     4501 tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg
     4561 ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgttct
     4621 ttcctgcgtt atcccctgat tctgtggata accgtattac cgcctttgag tgagctgata
     4681 ccgctcgccg cagccgaacg accgagcgca gcgagtcagt gagcgaggaa gcggaagagc
     4741 gcccaatacg caaaccgcct ctccccgcgc gttggccgat tcattaatgc agctggcacg
     4801 acaggtttcc cgactggaaa gcgggcagtg agcgcaacgc aattaatgtg agttagctca
     4861 ctcattaggc accccaggct ttacacttta tgcttccggc tcgtatgttg tgtggctcga
     4921 gtttactccc tatcagtgat agagaacgta tgtcgagttt actccctatc agtgatagag
     4981 aacgatgtcg agtttactcc ctatcagtga tagagaacgt atgtcgagtt tactccctat
     5041 cagtgataga gaacgtatgt cgagtttact ccctatcagt gatagagaac gtatgtcgag
     5101 tttatcccta tcagtgatag agaacgtatg tcgagtttac tccctatcag tgatagagaa
     5161 cgtatgtcga ggtaggcgtg tacggtggga ggcctatata agcagagctc gtttagtgaa
     5221 ccgtcagatc
//
