LOCUS       sequence-174393-        8654 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  HR Donor for N-terminal HA-mEOS3.2-TRF2 fusion at endogenous TRF2 
            locus in human cells..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8654)
  AUTHORS   Schmidt JC, Zaug AJ, Cech TR
  TITLE     Live Cell Imaging Reveals the Dynamics of Telomerase Recruitment to 
            Telomeres.
  JOURNAL   Cell. 2016 Aug 25;166(5):1188-1197.e9. doi: 
            10.1016/j.cell.2016.07.033. Epub 2016 Aug 11.
  PUBMED    27523609
REFERENCE   2  (bases 1 to 8654)
  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.cell.2016.07.033"; journalName: "Cell"; date: 
            "2016-08-25- 25"; volume: "166"; issue: "5"; pages: "1188-1197.e9"
COMMENT     Sequence Label: HA-mEOS3.2-TRF2 HR Donor
FEATURES             Location/Qualifiers
     source          1..8654
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      2..456
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(89..108)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     298..319
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        482..586
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(587..655,656..1447)
                     /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(805..824)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1618..2206
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2107..2126
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2349..2366
                     /label=L4440
                     /note="L4440 vector, forward primer"
     mobile_element  2500..2724
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7R
                     /note="mini-Tn7 element (right end of the Tn7 transposon)"
     CDS             complement(2791..3324)
                     /codon_start=1
                     /gene="aacC1"
                     /product="gentamycin acetyltransferase"
                     /label=GmR
                     /note="confers resistance to gentamycin"
                     /translation="MLRSSNDVTQQGSRPKTKLGGSSMGIIRTCRLGPDQVKSMRAALD
                     LFGREFGDVATYSQHQPDSDYLGNLLRSKTFIALAAFDQEAVVGALAAYVLPKFEQPRS
                     EIYIYDLAVSGEHRRQGIATALINLLKHEANALGAYVIYVQADYGDDPAVALYTKLGIR
                     EEVMHFDIDPSTAT"
     promoter        complement(join(3494..3499,3500..3516,3517..3522))
                     /gene="intI1 (promoter lies within the coding sequence)"
                     /label=Pc promoter
                     /note="class 1 integron promoter"
     promoter        3874..3965
                     /gene="polh from Autographa californica"
                     /label=polyhedrin promoter
                     /note="promoter for the baculovirus polyhedrin gene"
     primer_bind     3890..3907
                     /label=Polyhedrin-F
                     /note="Polyhedrin promoter, forward primer"
     CDS             4026..4043
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             4050..5147
                     /codon_start=1
                     /gene="malE (mutated)"
                     /product="maltose binding protein from E. coli"
                     /label=MBP
                     /note="This version of the gene does not encode a signal 
                     sequence, so MBP will remain in the cytosol."
                     /translation="KTEEGKLVIWINGDKGYNGLAEVGKKFEKDTGIKVTVEHPDKLEE
                     KFPQVAATGDGPDIIFWAHDRFGGYAQSGLLAEITPDKAFQDKLYPFTWDAVRYNGKLI
                     AYPIAVEALSLIYNKDLLPNPPKTWEEIPALDKELKAKGKSALMFNLQEPYFTWPLIAA
                     DGGYAFKYENGKYDIKDVGVDNAGAKAGLTFLVDLIKNKHMNADTDYSIAEAAFNKGET
                     AMTINGPWAWSNIDTSKVNYGVTVLPTFKGQPSKPFVGVLSAGINAASPNKELAKEFLE
                     NYLLTDEGLEAVNKDKPLGAVALKSYEEELAKDPRIAATMENAQKGEIMPNIPQMSAFW
                     YAVRTAVINAASGRQTVDEALKDAQT"
     regulatory      5004..5013
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     5121..5144
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     CDS             5175..5198
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             6570..6596
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             6597..7274
                     /codon_start=1
                     /product="improved monomeric variant of green-to-red 
                     photoswitchable fluorescent protein EosFP (Zhang et al., 
                     2012)"
                     /label=mEos3.2
                     /translation="MSAIKPDMKIKLRMEGNVNGHHFVIDGDGTGKPFEGKQSMDLEVK
                     EGGPLPFAFDILTTAFHYGNRVFAKYPDNIQDYFKQSFPKGYSWERSLTFEDGGICNAR
                     NDITMEGDTFYNKVRFYGTNFPANGPVMQKKTLKWEPSTEKMYVRDGVLTGDIEMALLL
                     EGNAHYRCDFRTTYKAKEKGVKLPGAHFVDHCIEILSHDKDYNKVKLYEHAVAHSGLPD
                     NARR"
     polyA_signal    8144..8278
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(8181..8200)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     8235..8254
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     mobile_element  8307..8472
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7L
                     /note="mini-Tn7 element (left end of the Tn7 transposon)"
ORIGIN
        1 gacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc
       61 gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc ctttctcgcc
      121 acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt
      181 agtgctttac ggcacctcga ccccaaaaaa cttgattagg gtgatggttc acgtagtggg
      241 ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt cttaatagtg
      301 gactcttgtt ccaaactgga acaacactca accctatctc ggtctattct tttgatttat
      361 aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa caaaaattta
      421 acgcgaattt taacaaaata ttaacgctta caatttaggt ggcacttttc ggggaaatgt
      481 gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc cgctcatgag
      541 acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtatga gtattcaaca
      601 tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt ttgctcaccc
      661 agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcacgag tgggttacat
      721 cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag aacgttttcc
      781 aatgatgagc acttttaaag ttctgctatg tggcgcggta ttatcccgta ttgacgccgg
      841 gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg agtactcacc
      901 agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca gtgctgccat
      961 aaccatgagt gataacactg cggccaactt acttctgaca acgatcggag gaccgaagga
     1021 gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc gttgggaacc
     1081 ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg tagcaatggc
     1141 aacaacgttg cgcaaactat taactggcga actacttact ctagcttccc ggcaacaatt
     1201 aatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg cccttccggc
     1261 tggctggttt attgctgata aatctggagc cggtgagcgt gggtctcgcg gtatcattgc
     1321 agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga cggggagtca
     1381 ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac tgattaagca
     1441 ttggtaactg tcagaccaag tttactcata tatactttag attgatttaa aacttcattt
     1501 ttaatttaaa aggatctagg tgaagatcct ttttgataat ctcatgacca aaatccctta
     1561 acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg
     1621 agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac cgctaccagc
     1681 ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa ctggcttcag
     1741 cagagcgcag ataccaaata ctgttcttct agtgtagccg tagttaggcc accacttcaa
     1801 gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag tggctgctgc
     1861 cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac cggataaggc
     1921 gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc gaacgaccta
     1981 caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc ccgaagggag
     2041 aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca cgagggagct
     2101 tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc tctgacttga
     2161 gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc
     2221 ggccttttta cggttcctgg ccttttgctc acatgttctt tcctgcgtta tcccctgatt
     2281 ctgtggataa ccgtattacc gcctttgagt gagctgatac cgctcgccgc agccgaacga
     2341 ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg cctgatgcgg tattttctcc
     2401 ttacgcatct gtgcggtatt tcacaccgca tagaccagcc gcgtaacctg gcaaaatcgg
     2461 ttacggttga gtaataaatg gatgccctgc gtaagcgggt gtgggcggac aataaagtct
     2521 taaactgaac aaaatagatc taaactatga caataaagtc ttaaactaga cagaatagtt
     2581 gtaaactgaa atcagtccag ttatgctgtg aaaaagcata ctggactttt gttatggcta
     2641 aagcaaactc ttcattttct gaagtgcaaa ttgcccgtcg tattaaagag gggcgtggcc
     2701 aagggcatgg taaagactat attcgcggcg ttgtgacaat ttaccgaaca actccgcggc
     2761 cgggaagccg atctcggctt gaacgaattg ttaggtggcg gtacttgggt cgatatcaaa
     2821 gtgcatcact tcttcccgta tgcccaactt tgtatagaga gccactgcgg gatcgtcacc
     2881 gtaatctgct tgcacgtaga tcacataagc accaagcgcg ttggcctcat gcttgaggag
     2941 attgatgagc gcggtggcaa tgccctgcct ccggtgctcg ccggagactg cgagatcata
     3001 gatatagatc tcactacgcg gctgctcaaa cttgggcaga acgtaagccg cgagagcgcc
     3061 aacaaccgct tcttggtcga aggcagcaag cgcgatgaat gtcttactac ggagcaagtt
     3121 cccgaggtaa tcggagtccg gctgatgttg ggagtaggtg gctacgtctc cgaactcacg
     3181 accgaaaaga tcaagagcag cccgcatgga tttgacttgg tcagggccga gcctacatgt
     3241 gcgaatgatg cccatacttg agccacctaa ctttgtttta gggcgactgc cctgctgcgt
     3301 aacatcgttg ctgctgcgta acatcaaaca tcgacccacg gcgtaacgcg cttgctgctt
     3361 ggatgcccga ggcatagact gtacaaaaaa acagtcataa caagccatga aaaccgccac
     3421 tgcgccgtta ccaccgctgc gttcggtcaa ggttctggac cagttgcgtg agcgcatacg
     3481 ctacttgcat tacagtttac gaaccgaaca ggcttatgtc aactgggttc gtgccttcat
     3541 ccgtttccac ggtgtgcgtc acccggcaac cttgggcagc agcgaagtcg aggcatttct
     3601 gtcctggctg gcgaacgagc gcaaggtttc ggtctccacg catcgtcagg cattggcggc
     3661 cttgctgttc ttctacggca aggtgctgtg cacggatctg ccctggcttc aggagatcgg
     3721 aagacctcgg ccgtcgcggc gcttgccggt ggtgctgacc ccggatgaag tggttcgcat
     3781 cctcggtttt ctggaaggcg agcatcgttt gttcgcccag gactctagct atagttctag
     3841 tggttggcta cgtatactcc ggaatattaa tagatcatgg agataattaa aatgataacc
     3901 atctcgcaaa taaataagta ttttactgtt ttcgtaacag ttttgtaata aaaaaaccta
     3961 taaatattcc ggattattca taccgtccca ccatcgggcg cggatctcgg tccgaaacca
     4021 tgggtcatca ccatcatcat cacggtacca aaactgaaga aggtaaactg gtaatctgga
     4081 ttaacggcga taaaggctat aacggtctcg ctgaagtcgg taagaaattc gagaaagata
     4141 ccggaattaa agtcaccgtt gagcatccgg ataaactgga agagaaattc ccacaggttg
     4201 cggcaactgg cgatggccct gacattatct tctgggcaca cgaccgcttt ggtggctacg
     4261 ctcaatctgg cctgttggct gaaatcaccc cggacaaagc gttccaggac aagctgtatc
     4321 cgtttacctg ggatgccgta cgttacaacg gcaagctgat tgcttacccg atcgctgttg
     4381 aagcgttatc gctgatttat aacaaagatc tgctgccgaa cccgccaaaa acctgggaag
     4441 agatcccggc gctggataaa gaactgaaag cgaaaggtaa gagcgcgctg atgttcaacc
     4501 tgcaagaacc gtacttcacc tggccgctga ttgctgctga cgggggttat gcgttcaagt
     4561 atgaaaacgg caagtacgac attaaagacg tgggcgtgga taacgctggc gcgaaagcgg
     4621 gtctgacctt cctggttgac ctgattaaaa acaaacacat gaatgcagac accgattact
     4681 ccatcgcaga agctgccttt aataaaggcg aaacagcgat gaccatcaac ggcccgtggg
     4741 catggtccaa catcgacacc agcaaagtga attatggtgt aacggtactg ccgaccttca
     4801 agggtcaacc atccaaaccg ttcgttggcg tgctgagcgc aggtattaac gccgccagtc
     4861 cgaacaaaga gctggcaaaa gagttcctcg aaaactatct gctgactgat gaaggtctgg
     4921 aagcggttaa taaagacaaa ccgctgggtg ccgtagcgct gaagtcttac gaggaagagt
     4981 tggcgaaaga tccacgtatt gccgccacca tggaaaacgc ccagaaaggt gaaatcatgc
     5041 cgaacatccc gcagatgtcc gctttctggt atgccgtgcg tactgcggtg atcaacgccg
     5101 ccagcggtcg tcagactgtc gatgaagccc tgaaagacgc gcagactggt accgattacg
     5161 atatcccaac gaccctggaa gttctgttcc aggggcccgg gtccatggag acgtgaaaat
     5221 aaaaatagcg tctcagagca taatctagag cctgcagtct cgaggcatgc ggtaccaagc
     5281 ttgtcgagaa gtactagagg atcataatca gccataccac atttgtagag gttttacttg
     5341 ctttaaaaaa cctcccacac ctccccctga acctgaaaca taaaatgaat gcaattgttg
     5401 ttgttggcca cctgcacttc tgagttgtct ttgagtggcc ccaaactgga tctttgcagg
     5461 ttccagacta gtggggctgg taggacaatc tttgaagcta gaccagttac tgggcctctc
     5521 aggaagaggc acccacaacc accatcctga actaaggtct gagctggagg ttctccattt
     5581 ggtcaagtgc cagttggtga attccaagag tatcagagat atctgaaagt gactcagacc
     5641 ctctatggga ctatgggtgg tccttttgct tatttttcta attgttctac aacaaaatgt
     5701 gttatttttg tgattacaaa gagtatatgt ctagcggctg ggttgccagg tgcattctat
     5761 ttattggtat cacactggtc tttgctttat aattacagta tttgttaaaa tacaaatttg
     5821 tttaaagact ttcctctgtg tatgctctct tccacaattt tcaaagtgtt taataaggtg
     5881 cacatatttg gttttctttt ttttcttttc ttttttttga gacggagttt cgctcttgtt
     5941 gcccagactg gagtgcaatg gcgcgatctc ggctcaccgc aacctccgcc tcgcgggttc
     6001 aagcgagtct cctgcctcag cctcctgagt agctaggatt acaggcatgc gccaccacgt
     6061 ctggctaatt ttgtattttt agtagagatg gggtttctcc atgttggtca ggctggtctc
     6121 gaactcccga cctcaggtga tccacccgcc tcagcctccc aaagtgctgg gattacaggc
     6181 gtagagccac cgatcccggc ctgtttttca gtctctaata cactgttccg cgtccctagc
     6241 atagtagctg ttttctgtaa attcggagcg aagttgactg caaaacccgc cgggttcacc
     6301 ggcctgggcg gtgtggactt caagccccag gaggcattgc ggccggcaca tcgggaacta
     6361 cggcgtctga gaagggggcc gggccctccg gtcctttccc tcccagaagc ccgcgggcgg
     6421 ctcggaacgc tgtttctatc atggccgcgg gagccgggac ggcgggcccc gcttccggcc
     6481 cgggcgtcgt gcgtgaccca gcggcgtcac agccgaggaa gcggcccggc cgggagggcg
     6541 gggagggcgc gcggcgatcg gacacgatgt acccatacga tgttccagat tacgctatga
     6601 gtgcgattaa gccagacatg aagatcaaac tccgtatgga aggcaacgta aacgggcacc
     6661 actttgtgat cgacggagat ggtacaggca agccttttga gggaaaacag agtatggatc
     6721 ttgaagtcaa agagggcgga cctctgcctt ttgcctttga tatcctgacc actgcattcc
     6781 attacggcaa cagggtattc gccaaatatc cagacaacat acaagactat tttaagcagt
     6841 cgtttcctaa ggggtattcg tgggaacgaa gcttgacttt cgaagacggg ggcatttgca
     6901 acgccagaaa cgacataaca atggaagggg acactttcta taataaagtt cgattttatg
     6961 gtaccaactt tcccgccaat ggtccagtta tgcagaagaa gacgctgaaa tgggagccct
     7021 ccactgagaa aatgtatgtg cgtgatggag tgctgacggg tgatattgag atggctttgt
     7081 tgcttgaagg aaatgcccat taccgatgtg acttcagaac tacttacaaa gctaaggaga
     7141 agggtgtcaa gttaccaggc gcccactttg tggaccactg cattgagatt ttaagccatg
     7201 acaaagatta caacaaggtt aagctgtatg agcatgctgt tgctcattct ggattgcctg
     7261 acaatgccag acgaggatcc atggcgggag gaggcgggag tagcgacggc agcgggcggg
     7321 cagctggcag gcgggcgtcc cgcagtagcg ggcgggcccg gcgggggcgc cacgagccgg
     7381 ggctgggggg cccggcggag cgcggcgcgg gggaggcacg gctggaagag gcagtcaatc
     7441 gctgggtgct caagttctac ttccacgagg cgctgcgggc ctttcggggt agccggtacg
     7501 gggacttcag acagatccgg gacatcatgc agggtgaggg ccgggccggg ggaggggggt
     7561 tggagcgccg gggaaggggg tccgggggac cgggagtcgg gactgggggt cgcccagcca
     7621 acgatgtctt ctctcccagc tttgcttgtc aggcccttgg ggaaggagca caccgtgtcc
     7681 cgattgctgc gggttatgca gtgtctgtcg cggattgaag aaggggaaaa tttaggtatg
     7741 gctatttttg tgttcttctc tgggcttact tgcgtagggt tttgttttaa aacttatatc
     7801 tagaactgaa ggattccagg ttaaagtgcg atagtttcca tcagagtggt ctttttcatt
     7861 ctacgatggg tctgttttct ggctcgactg gctctagttt tctccggtta aggttgtctt
     7921 ggctgctttt ctgtgttttt tttaaatctt ctcccacctg ttaattgctc accccaactt
     7981 tgctcctgtc taaagtgagt tatgttcata tggacatctt ttttccccca tggtagagag
     8041 tttttcagag tggaagtact ctgacatttt tcccctctga atttaacttg aaagctattt
     8101 agctgttgtc agtttcttgc cactcgtatc tgaggaggat gggaacttgt ttattgcagc
     8161 ttataatggt tacaaataaa gcaatagcat cacaaatttc acaaataaag catttttttc
     8221 actgcattct agttgtggtt tgtccaaact catcaatgta tcttatcatg tctggatctg
     8281 atcactgata tcgcctagga gatccgaacc agataagtga aatctagttc caaactattt
     8341 tgtcattttt aattttcgta ttagcttacg acgctacacc cagttcccat ctattttgtc
     8401 actcttccct aaataatcct taaaaactcc atttccaccc ctcccagttc ccaactattt
     8461 tgtccgccca cagcggggca tttttcttcc tgttatgttt ttaatcaaac atcctgccaa
     8521 ctccatgtga caaaccgtca tcttcggcta ctttttctct gtcacagaat gaaaattttt
     8581 ctgtcatctc ttcgttatta atgtttgtaa ttgactgaat atcaacgctt atttgcagcc
     8641 tgaatggcga atgg
//
