LOCUS       sequence-448157-        4892 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Generates TELO2 dTAG N terminal knock-in cell lines.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4892)
  AUTHORS   Huang M, Yao F, Nie L, Wang C, Su D, Zhang H, Li S, Tang M, Feng X, 
            Yu B, Chen Z, Wang S, Yin L, Mou L, Hart T, Chen J
  TITLE     FACS-based genome-wide CRISPR screens define key regulators of DNA 
            damage signaling pathways.
  JOURNAL   Mol Cell. 2023 Aug 3;83(15):2810-2828.e6. doi: 
            10.1016/j.molcel.2023.07.004.
  PUBMED    37541219
REFERENCE   2  (bases 1 to 4892)
  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: 
            "10.1016/j.molcel.2023.07"; journalName: "Mol Cell"; date: 
            "2023-08-3- 3"; volume: "83"; issue: "15"; pages: "2810-2828.e6"
COMMENT     Sequence Label: pUC19_TELO2_dTAG
FEATURES             Location/Qualifiers
     source          1..4892
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     118..137
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     371..388
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    505..526
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(541..546,547..564,565..571)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    579..595
                     /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     584..606
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     603..619
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     603..619
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             615..938
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITPSLHACRSTLEDPRVPSSNSLAVVLQRRDWENPGVTQLNR
                     LAAHPPFASWRNSEEARTDRPSQQLRSLNGEWRLMRYFLLTHLCGISHRIWCTLSTICS
                     DAA"
     misc_feature    632..688
                     /label=MCS
                     /note="pUC18/19 multiple cloning site"
     primer_bind     complement(689..706)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(689..705)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(698..720)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(914..933)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     CDS             1527..1922
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     CDS             1932..1988
                     /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             1989..2015
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /note="This is a variant of the standard HA tag, which has 
                     sequence YPYDVPDYA. Some anti-HA antibodies may not 
                     recognize this variant."
                     /translation="YPYDVPDYA"
     CDS             1989..2015
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             2019..2045
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             2055..2375
                     /codon_start=1
                     /product="F36V mutant of FK506-binding protein FKBP12"
                     /label=FKBP-V
                     /note="binds synthetic ligands that do not bind wild-type 
                     FKBP"
                     /translation="GVQVETISPGDGRTFPKRGQTCVVHYTGMLEDGKKVDSSRDRNKP
                     FKFMLGKQEVIRGWEEGVAQMSVGQRAKLTISPDYAYGATGHPGIIPPHATLVFDVELL
                     KLE"
     primer_bind     3239..3261
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3299..3317)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3385..3489
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3490..3558,3559..4350)
                     /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(3708..3727)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      join(4521..4892,1..217)
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga
       61 caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg
      121 aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt
      181 tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt
      241 acggttcctg gccttttgct ggccttttgc tcacatgttc tttcctgcgt tatcccctga
      301 ttctgtggat aaccgtatta ccgcctttga gtgagctgat accgctcgcc gcagccgaac
      361 gaccgagcgc agcgagtcag tgagcgagga agcggaagag cgcccaatac gcaaaccgcc
      421 tctccccgcg cgttggccga ttcattaatg cagctggcac gacaggtttc ccgactggaa
      481 agcgggcagt gagcgcaacg caattaatgt gagttagctc actcattagg caccccaggc
      541 tttacacttt atgcttccgg ctcgtatgtt gtgtggaatt gtgagcggat aacaatttca
      601 cacaggaaac agctatgacc atgattacgc caagcttgca tgcctgcagg tcgactctag
      661 aggatccccg ggtaccgagc tcgaattcac tggccgtcgt tttacaacgt cgtgactggg
      721 aaaaccctgg cgttacccaa cttaatcgcc ttgcagcaca tccccctttc gccagctggc
      781 gtaatagcga agaggcccgc accgatcgcc cttcccaaca gttgcgcagc ctgaatggcg
      841 aatggcgcct gatgcggtat tttctcctta cgcatctgtg cggtatttca caccgcatat
      901 ggtgcactct cagtacaatc tgctctgatg ccgcatagtt aagccagccc cgacacccgc
      961 caacacccgc tgacgcgccc tgacgggctt gtctgctccc ggcatccgct tacagacaag
     1021 ctgtgaccgt ctccgtaagc tgttggtttc taaggggcgg aggaatggcg actggagcca
     1081 cctctcaccg ctcagggagc cgggagggcc ggcagctccc gcgggccgag tttcccgcct
     1141 tgggtgcgag gacgcgtggg gccgcgctgt gcccggggaa cactcaccag catctctagc
     1201 ctccacctcc ctctcgcgcc agtggcactc ccggttgaga caactgagaa tgttccctgg
     1261 gcacagtggc ccgggttgag aagccctgca gtggctggta gcgttggggt ccgagcggag
     1321 gagcgaactc tgggtggaaa ccggcaggca ctggagggga aggaggcggg acagggttgg
     1381 gtgggaccag ggttgagggg tgtggggtct cggggcgctc accgaggggc ttcctgagct
     1441 tgtgtggatt atttccgtgc cccaagctga gccctgtgtc catgtcacag gtcgtcttcc
     1501 cgtgacgccc agatctgtgc tactggatgg ccaagccttt gtctcaagaa gaatccaccc
     1561 tcattgaaag agcaacggct acaatcaaca gcatccccat ctctgaagac tacagcgtcg
     1621 ccagcgcagc tctctctagc gacggccgca tcttcactgg tgtcaatgta tatcatttta
     1681 ctgggggacc ttgtgcagaa ctcgtggtgc tgggcactgc tgctgctgcg gcagctggca
     1741 acctgacttg tatcgtcgcg atcggaaatg agaacagggg catcttgagc ccctgcggac
     1801 ggtgccgaca ggtgcttctc gatctgcatc ctgggatcaa agccatagtg aaggacagtg
     1861 atggacagcc gacggcagtt gggattcgtg aattgctgcc ctctggttat gtgtgggagg
     1921 gcggaagcgg agctactaac ttcagcctgc tgaagcaggc tggagacgtg gaggagaacc
     1981 ctggacctta cccctacgac gtgcccgact acgccggcta tccgtatgat gtcccggact
     2041 atgcaggaag cggaggagtg caggtggaaa ccatctcccc aggagacggg cgcaccttcc
     2101 ccaagcgcgg ccagacctgc gtggtgcact acaccgggat gcttgaagat ggaaagaaag
     2161 ttgattcctc ccgggacaga aacaagccct ttaagtttat gctaggcaag caggaggtga
     2221 tccgaggctg ggaagaaggg gttgcccaga tgagtgtggg tcagagagcc aaactgacta
     2281 tatctccaga ttatgcctat ggtgccactg ggcacccagg catcatccca ccacatgcca
     2341 ctctcgtctt cgatgtggag cttctaaaac tggaaggtgg cggtggctcg ggcggtggtg
     2401 ggtcgggtgg cggcggatct atggagccag caccctcaga ggttcgactc gccgtccggg
     2461 aagccattca tgccctctcg tcttcggagg atggcggcca catcttctgc accctggagt
     2521 ccctgaagcg gtatctcggt gagatggagc ctccagcgct cccgagggag aaggaggagt
     2581 ttgcctcggc ccacttctcg cctgtcctca gatgtcttgc cagcaggctg agcccagcct
     2641 ggctggagct gctgccccat ggccgcctgg aggagctgtg ggccagcttc ttcctggagg
     2701 gcccggcgga ccaagccttc ctggtgttga tggagaccat cgagggtgct gcggggtgag
     2761 tgggctgggc ccatcctggg gttgccggta gcctcagaag tgatgagagt ggcttgaagg
     2821 actggaccaa gagcctctct agtccctgtg aggggctaga gagagagcct gctcctggct
     2881 gaacccctga acagaagaag cggtctgtgt ctgtctcctt tgcgacggga ggcacctgct
     2941 gtgtctcaca aagtccccca cttgctcccc gtccggctgt gtcagagagg gatggggtgg
     3001 gagtgttcac atcccaggcg gcagaggcag cccgtcagct ggggacgttc caggttttcc
     3061 agggaagcac agctgtcatc actgacacgt gtcccatgtg gactcccagc cccagccggc
     3121 tcccaggctc agcctcagat cacactgcag tccttgccgg gacactgcat cttgagaagc
     3181 tgggttttca gaggttgctg tgattaaagc aagcactgag ttatccgcgt cagcagcacc
     3241 gggagctgca tgtgtcagag gttttcaccg tcatcaccga aacgcgcgag acgaaagggc
     3301 ctcgtgatac gcctattttt ataggttaat gtcatgataa taatggtttc ttagacgtca
     3361 ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat
     3421 tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa
     3481 aggaagagta tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt
     3541 tgccttcctg tttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag
     3601 ttgggtgcac gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt
     3661 tttcgccccg aagaacgttt tccaatgatg agcactttta aagttctgct atgtggcgcg
     3721 gtattatccc gtattgacgc cgggcaagag caactcggtc gccgcataca ctattctcag
     3781 aatgacttgg ttgagtactc accagtcaca gaaaagcatc ttacggatgg catgacagta
     3841 agagaattat gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg
     3901 acaacgatcg gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta
     3961 actcgccttg atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac
     4021 accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg cgaactactt
     4081 actctagctt cccggcaaca attaatagac tggatggagg cggataaagt tgcaggacca
     4141 cttctgcgct cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag
     4201 cgtgggtctc gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta
     4261 gttatctaca cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag
     4321 ataggtgcct cactgattaa gcattggtaa ctgtcagacc aagtttactc atatatactt
     4381 tagattgatt taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat
     4441 aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta
     4501 gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa
     4561 acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt
     4621 tttccgaagg taactggctt cagcagagcg cagataccaa atactgttct tctagtgtag
     4681 ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta
     4741 atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca
     4801 agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag
     4861 cccagcttgg agcgaacgac ctacaccgaa ct
//
