LOCUS       sequence-224968-        5072 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Repair template for the TLR reporter with 10 UAS binding sites for 
            yeast Gal4.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5072)
  AUTHORS   Bashir S, Dang T, Rossius J, Wolf J, Kuhn R
  TITLE     Enhancement of CRISPR-Cas9 induced precise gene editing by targeting
            histone H2A-K15 ubiquitination.
  JOURNAL   BMC Biotechnol. 2020 Oct 23;20(1):57. doi: 
            10.1186/s12896-020-00650-x.
  PUBMED    33097066
REFERENCE   2  (bases 1 to 5072)
  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: "BMC 
            Biotechnol."; date: "2020-10-23"; volume: "20(1):57. doi"; pages: " 
            10.1186/s12896-020-00650-x"
COMMENT     Sequence Label: pTLR-donor-UAS
FEATURES             Location/Qualifiers
     source          1..5072
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    52..146
                     /label=5X UAS
                     /bound_moiety="GAL4"
                     /note="five tandem copies of the ""ScaI site"" 17-mer 
                     CGGAGTACTGTCCTCCG, an upstream activating sequence (UAS) 
                     that efficiently binds yeast Gal4 (Webster et al., 1988; 
                     Pfeiffer et al., 2010)"
     protein_bind    199..293
                     /label=5X UAS
                     /bound_moiety="GAL4"
                     /note="five tandem copies of the ""ScaI site"" 17-mer 
                     CGGAGTACTGTCCTCCG, an upstream activating sequence (UAS) 
                     that efficiently binds yeast Gal4 (Webster et al., 1988; 
                     Pfeiffer et al., 2010)"
     CDS             join(353..355,356..1069)
                     /codon_start=1
                     /product="yellow fluorescent protein (YFP) with fast and 
                     efficient maturation (Nagai et al., 2002)"
                     /label=Venus
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     LICTTGKLPVPWPTLVTTLGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIKA
                     NFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(395..416)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(656..675)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1003..1024
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             1078..1134
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             1135..1848
                     /codon_start=1
                     /product="monomeric derivative of red fluorescent protein 
                     from Entacmaea quadricolor (Merzlyak et al., 2007)"
                     /label=TagRFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELIKENMHMKLYMEGTVNNHHFKCTSEGEGKPYEGTQTM
                     RIKVVEGGPLPFAFDILATSFMYGSRTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGG
                     VLTATQDTSLQDGCLIYNVKIRGVNFPSNGPVMQKKTLGWEANTEMLYPADGGLEGRSD
                     MALKLVGGGHLICNFKTTYRSKKPAKNLKMPGVYYVDHRLERIKEADKETYVEQHEVAV
                     ARYCDLPSKLGHKLN"
     primer_bind     complement(1874..1890)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1904..1921)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1910..2134
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(2228..2247)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2229..2247)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2257..2274)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2257..2273)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2266..2288)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      2415..2870
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2502..2521)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2712..2733
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        2896..3000
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3001..3069,3070..3861)
                     /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(3219..3238)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4032..4620
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4521..4540
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4774..4791
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    4908..4929
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(4944..4949,4950..4967,4968..4974)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4982..4998
                     /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)."
     primer_bind     4987..5009
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     5006..5022
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5006..5022
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     5041..5061
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        5043..5061
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
ORIGIN
        1 gggtaccggg ccccccctcg aggtcgaccg caagcttgca tgcctgcagg tcggagtact
       61 gtcctccgag cggagtactg tcctccgagc ggagtactgt cctccgagcg gagtactgtc
      121 ctccgagcgg agtactgtcc tccgagcgga gactctagcg ctagcgcatg tcgaccgcaa
      181 gcttgcatgc ctgcaggtcg gagtactgtc ctccgagcgg agtactgtcc tccgagcgga
      241 gtactgtcct ccgagcggag tactgtcctc cgagcggagt actgtcctcc gagcggagac
      301 tctagcgcta gcgcatgtcg acggtatcga taagcttgat taagccgcca ccgtgagcaa
      361 gggcgaggag ctgttcaccg gggtggtgcc catcctggtc gagctggacg gcgacgtaaa
      421 cggccacaag ttcagcgtgt ccggcgaggg cgagggcgat gccacctacg gcaagctgac
      481 cctgaagctg atctgcacca ccggcaagct gcccgtgccc tggcccaccc tcgtgaccac
      541 cctgggctac ggcctgcagt gcttcgcccg ctaccccgac cacatgaagc agcacgactt
      601 cttcaagtcc gccatgcccg aaggctacgt ccaggagcgc accatcttct tcaaggacga
      661 cggcaactac aagacccgcg ccgaggtgaa gttcgagggc gacaccctgg tgaaccgcat
      721 cgagctgaag ggcatcgact tcaaggagga cggcaacatc ctggggcaca agctggagta
      781 caactacaac agccacaacg tctatatcac cgccgacaag cagaagaacg gcatcaaggc
      841 caacttcaag atccgccaca acatcgagga cggcggcgtg cagctcgccg accactacca
      901 gcagaacacc cccatcggcg acggccccgt gctgctgccc gacaaccact acctgagcta
      961 ccagtccgcc ctgagcaaag accccaacga gaagcgcgat cacatggtcc tgctggagtt
     1021 cgtgaccgcc gccgggatca ctctcggcat ggacgagctg tacaagtaga cgcgttggcc
     1081 acgaacttct ctctgttaaa gcaagcagga gatgttgaag aaaaccccgg gcctatggtg
     1141 tctaagggcg aagagctgat taaggagaac atgcacatga agctgtacat ggagggcacc
     1201 gtgaacaacc accacttcaa gtgcacatcc gagggcgaag gcaagcccta cgagggcacc
     1261 cagaccatga gaatcaaggt ggtcgagggc ggccctctcc ccttcgcctt cgacatcctg
     1321 gctaccagct tcatgtacgg cagcagaacc ttcatcaacc acacccaggg catccccgac
     1381 ttctttaagc agtccttccc tgagggcttc acatgggaga gagtcaccac atacgaagac
     1441 gggggcgtgc tgaccgctac ccaggacacc agcctccagg acggctgcct catctacaac
     1501 gtcaagatca gaggggtgaa cttcccatcc aacggccctg tgatgcagaa gaaaacactc
     1561 ggctgggagg ccaacaccga gatgctgtac cccgctgacg gcggcctgga aggcagaagc
     1621 gacatggccc tgaagctcgt gggcgggggc cacctgatct gcaacttcaa gaccacatac
     1681 agatccaaga aacccgctaa gaacctcaag atgcccggcg tctactatgt ggaccacaga
     1741 ctggaaagaa tcaaggaggc cgacaaagag acctacgtcg agcagcacga ggtggctgtg
     1801 gccagatact gcgacctccc tagcaaactg gggcacaaac ttaattgagc gatcgcacgc
     1861 gtaaatgatt gcagatccac tagttctaga gctcgctgat cagcctcgac tgtgccttct
     1921 agttgccagc catctgttgt ttgcccctcc cccgtgcctt ccttgaccct ggaaggtgcc
     1981 actcccactg tcctttccta ataaaatgag gaaattgcat cgcattgtct gagtaggtgt
     2041 cattctattc tggggggtgg ggtggggcag gacagcaagg gggaggattg ggaagacaat
     2101 agcaggcatg ctggggatgc ggtgggctct atggcttctg aggcggaaag aaccagctgg
     2161 ggctcgagat ccactagttc tagcctcgag gctagagcgg ccgccaccgc ggtggagctc
     2221 caattcgccc tatagtgagt cgtattacgc gcgctcactg gccgtcgttt tacaacgtcg
     2281 tgactgggaa aaccctggcg ttacccaact taatcgcctt gcagcacatc cccctttcgc
     2341 cagctggcgt aatagcgaag aggcccgcac cgatcgccct tcccaacagt tgcgcagcct
     2401 gaatggcgaa tgggacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac
     2461 gcgcagcgtg accgctacac ttgccagcgc cctagcgccc gctcctttcg ctttcttccc
     2521 ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt
     2581 agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt agggtgatgg
     2641 ttcacgtagt gggccatcgc cctgatagac ggtttttcgc cctttgacgt tggagtccac
     2701 gttctttaat agtggactct tgttccaaac tggaacaaca ctcaacccta tctcggtcta
     2761 ttcttttgat ttataaggga ttttgccgat ttcggcctat tggttaaaaa atgagctgat
     2821 ttaacaaaaa tttaacgcga attttaacaa aatattaacg cttacaattt aggtggcact
     2881 tttcggggaa atgtgcgcgg aacccctatt tgtttatttt tctaaataca ttcaaatatg
     2941 tatccgctca tgagacaata accctgataa atgcttcaat aatattgaaa aaggaagagt
     3001 atgagtattc aacatttccg tgtcgccctt attccctttt ttgcggcatt ttgccttcct
     3061 gtttttgctc acccagaaac gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca
     3121 cgagtgggtt acatcgaact ggatctcaac agcggtaaga tccttgagag ttttcgcccc
     3181 gaagaacgtt ttccaatgat gagcactttt aaagttctgc tatgtggcgc ggtattatcc
     3241 cgtattgacg ccgggcaaga gcaactcggt cgccgcatac actattctca gaatgacttg
     3301 gttgagtact caccagtcac agaaaagcat cttacggatg gcatgacagt aagagaatta
     3361 tgcagtgctg ccataaccat gagtgataac actgcggcca acttacttct gacaacgatc
     3421 ggaggaccga aggagctaac cgcttttttg cacaacatgg gggatcatgt aactcgcctt
     3481 gatcgttggg aaccggagct gaatgaagcc ataccaaacg acgagcgtga caccacgatg
     3541 cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg gcgaactact tactctagct
     3601 tcccggcaac aattaataga ctggatggag gcggataaag ttgcaggacc acttctgcgc
     3661 tcggcccttc cggctggctg gtttattgct gataaatctg gagccggtga gcgtgggtct
     3721 cgcggtatca ttgcagcact ggggccagat ggtaagccct cccgtatcgt agttatctac
     3781 acgacgggga gtcaggcaac tatggatgaa cgaaatagac agatcgctga gataggtgcc
     3841 tcactgatta agcattggta actgtcagac caagtttact catatatact ttagattgat
     3901 ttaaaacttc atttttaatt taaaaggatc taggtgaaga tcctttttga taatctcatg
     3961 accaaaatcc cttaacgtga gttttcgttc cactgagcgt cagaccccgt agaaaagatc
     4021 aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa
     4081 ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct ttttccgaag
     4141 gtaactggct tcagcagagc gcagatacca aatactgtcc ttctagtgta gccgtagtta
     4201 ggccaccact tcaagaactc tgtagcaccg cctacatacc tcgctctgct aatcctgtta
     4261 ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg ggttggactc aagacgatag
     4321 ttaccggata aggcgcagcg gtcgggctga acggggggtt cgtgcacaca gcccagcttg
     4381 gagcgaacga cctacaccga actgagatac ctacagcgtg agctatgaga aagcgccacg
     4441 cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg aacaggagag
     4501 cgcacgaggg agcttccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc
     4561 cacctctgac ttgagcgtcg atttttgtga tgctcgtcag gggggcggag cctatggaaa
     4621 aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg
     4681 ttctttcctg cgttatcccc tgattctgtg gataaccgta ttaccgcctt tgagtgagct
     4741 gataccgctc gccgcagccg aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa
     4801 gagcgcccaa tacgcaaacc gcctctcccc gcgcgttggc cgattcatta atgcagctgg
     4861 cacgacaggt ttcccgactg gaaagcgggc agtgagcgca acgcaattaa tgtgagttag
     4921 ctcactcatt aggcacccca ggctttacac tttatgcttc cggctcgtat gttgtgtgga
     4981 attgtgagcg gataacaatt tcacacagga aacagctatg accatgatta cgccaagcgc
     5041 gcaattaacc ctcactaaag ggaacaaaag ct
//
