LOCUS       sequence-73769-e        8345 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Left TALEN for PKP2 with NK.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8345)
  AUTHORS   Miyaoka Y, Chan AH, Judge LM, Yoo J, Huang M, Nguyen TD, Lizarraga 
            PP, So PL, Conklin BR
  TITLE     Isolation of single-base genome-edited human iPS cells without 
            antibiotic selection.
  JOURNAL   Nat Methods. 2014 Mar;11(3):291-3. doi: 10.1038/nmeth.2840. Epub 
            2014 Feb 9.
  PUBMED    24509632
REFERENCE   2  (bases 1 to 8345)
  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: 
            "10.1038/nmeth.2840"; journalName: "Nat Methods"; date: "2014-03"; 
            volume: "11"; issue: "3"; pages: "291-3"
COMMENT     Sequence Label: MR015-PKP2 Left TALEN_NK
FEATURES             Location/Qualifiers
     source          1..8345
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      901..910
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             910..975
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             982..1002
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(3939..3956)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3945..4169
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      4215..4643
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4302..4321)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4512..4533
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4652..4672)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4657..4986
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4837..4972
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4899..4918
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             5053..5847
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(5107..5126)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5717..5736
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    6021..6142
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6058..6077)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6112..6131
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(6191..6207)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6191..6207)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6204..6226)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6215..6231
                     /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(6239..6245,6246..6263,6264..6269))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6284..6305
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6422..6439)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6593..7178)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6673..6692)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7349..8140,8141..8209))
                     /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     7972..7991
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8210..8314)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc
      901 gccaccatgg actacaaaga ccatgacggt gattataaag atcatgacat cgattacaag
      961 gatgacgatg acaagatggc ccccaagaag aagaggaagg tgggcattca ccgcggggta
     1021 cctatggtgg acttgaggac actcggttat tcgcaacagc aacaggagaa aatcaagcct
     1081 aaggtcagga gcaccgtcgc gcaacaccac gaggcgcttg tggggcatgg cttcactcat
     1141 gcgcatattg tcgcgctttc acagcaccct gcggcgcttg ggacggtggc tgtcaaatac
     1201 caagatatga ttgcggccct gcccgaagcc acgcacgagg caattgtagg ggtcggtaaa
     1261 cagtggtcgg gagcgcgagc acttgaggcg ctgctgactg tggcgggtga gcttaggggg
     1321 cctccgctcc agctcgacac cgggcagctg ctgaagatcg cgaagagagg gggagtaaca
     1381 gcggtagagg cagtgcacgc ctggcgcaat gcgctcaccg gtgcccccct gaacctgacc
     1441 ccggaccaag tggtggctat cgccagcaac aagggcggca agcaagcgct cgaaacggtg
     1501 cagcggctgt tgccggtgct gtgccaggac catggcctga ccccggacca agtggtggct
     1561 atcgccagca acattggcgg caagcaagcg ctcgaaacgg tgcagcggct gttgccggtg
     1621 ctgtgccagg accatggcct gactccggac caagtggtgg ctatcgccag ccacgatggc
     1681 ggcaagcaag cgctcgaaac ggtgcagcgg ctgttgccgg tgctgtgcca ggaccatggc
     1741 ctgaccccgg accaagtggt ggctatcgcc agcaacaagg gcggcaagca agcgctcgaa
     1801 acggtgcagc ggctgttgcc ggtgctgtgc caggaccatg gcctgacccc ggaccaagtg
     1861 gtggctatcg ccagcaacaa gggcggcaag caagcgctcg aaacggtgca gcggctgttg
     1921 ccggtgctgt gccaggacca tggcctgacc ccggaccaag tggtggctat cgccagcaac
     1981 attggcggca agcaagcgct cgaaacggtg cagcggctgt tgccggtgct gtgccaggac
     2041 catggcctga ccccggacca agtggtggct atcgccagca acaagggcgg caagcaagcg
     2101 ctcgaaacgg tgcagcggct gttgccggtg ctgtgccagg accatggcct gaccccggac
     2161 caagtggtgg ctatcgccag caacattggc ggcaagcaag cgctcgaaac ggtgcagcgg
     2221 ctgttgccgg tgctgtgcca ggaccatggc ctgaccccgg accaagtggt ggctatcgcc
     2281 agcaacattg gcggcaagca agcgctcgaa acggtgcagc ggctgttgcc ggtgctgtgc
     2341 caggaccatg gcctgacccc ggaccaagtg gtggctatcg ccagcaacgg tggcggcaag
     2401 caagcgctcg aaacggtgca gcggctgttg ccggtgctgt gccaggacca tggcctgacc
     2461 ccggaccaag tggtggctat cgccagcaac attggcggca agcaagcgct cgaaacggtg
     2521 cagcggctgt tgccggtgct gtgccaggac catggcctga ccccggacca agtggtggct
     2581 atcgccagca acggtggcgg caagcaagcg ctcgaaacgg tgcagcggct gttgccggtg
     2641 ctgtgccagg accatggcct gactccggac caagtggtgg ctatcgccag ccacgatggc
     2701 ggcaagcaag cgctcgaaac ggtgcagcgg ctgttgccgg tgctgtgcca ggaccatggc
     2761 ctgaccccgg accaagtggt ggctatcgcc agcaacattg gcggcaagca agcgctcgaa
     2821 acggtgcagc ggctgttgcc ggtgctgtgc caggaccatg gcctgacccc ggaccaagtg
     2881 gtggctatcg ccagcaacgg tggcggcaag caagcgctcg aaacggtgca gcggctgttg
     2941 ccggtgctgt gccaggacca tggcctgact ccggaccaag tggtggctat cgccagccac
     3001 gatggcggca agcaagcgct cgaaacggtg cagcggctgt tgccggtgct gtgccaggac
     3061 catggcctga ccccggacca agtggtggct atcgccagca acattggcgg caagcaagcg
     3121 ctcgaaagca ttgtggccca gctgagccgg cctgatccgg cgttggccgc gttgaccaac
     3181 gaccatctgg tggcgttggc atgtcttggt ggacgacccg cgctcgatgc agtcaaaaag
     3241 ggtctgcctc atgctcccgc attgatcaaa agaaccaacc ggcggattcc cgagagaact
     3301 tcccatcgag tcgcgggatc ccagctggtg aagagcgagc tggaggagaa gaagtccgag
     3361 ctgcggcaca agctgaagta cgtgccccac gagtacatcg agctgatcga gatcgccagg
     3421 aacagcaccc aggaccgcat cctggagatg aaggtgatgg agttcttcat gaaggtgtac
     3481 ggctacaggg gaaagcacct gggcggaagc agaaagcctg acggcgccat ctatacagtg
     3541 ggcagcccca tcgattacgg cgtgatcgtg gacacaaagg cctacagcgg cggctacaat
     3601 ctgcctatcg gccaggccga cgagatgcag agatacgtgg aggagaacca gacccggaat
     3661 aagcacatca accccaacga gtggtggaag gtgtacccta gcagcgtgac cgagttcaag
     3721 ttcctgttcg tgagcggcca cttcaagggc aactacaagg cccagctgac caggctgaac
     3781 cacatcacca accgcaatgg cgccgtgctg agcgtggagg agctgctgat cggcggcgag
     3841 atgatcaaag ccggcaccct gacactggag gaggtgcggc gcaagttcaa caacggcgag
     3901 atcaacttct gataacttaa gtttaaaccg ctgatcagcc tcgactgtgc cttctagttg
     3961 ccagccatct gttgtttgcc cctcccccgt gccttccttg accctggaag gtgccactcc
     4021 cactgtcctt tcctaataaa atgaggaaat tgcatcgcat tgtctgagta ggtgtcattc
     4081 tattctgggg ggtggggtgg ggcaggacag caagggggag gattgggaag acaatagcag
     4141 gcatgctggg gatgcggtgg gctctatggc ttctgaggcg gaaagaacca gctggggctc
     4201 tagggggtat ccccacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac
     4261 gcgcagcgtg accgctacac ttgccagcgc cctagcgccc gctcctttcg ctttcttccc
     4321 ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt
     4381 agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt agggtgatgg
     4441 ttcacgtagt gggccatcgc cctgatagac ggtttttcgc cctttgacgt tggagtccac
     4501 gttctttaat agtggactct tgttccaaac tggaacaaca ctcaacccta tctcggtcta
     4561 ttcttttgat ttataaggga ttttgccgat ttcggcctat tggttaaaaa atgagctgat
     4621 ttaacaaaaa tttaacgcga attaattctg tggaatgtgt gtcagttagg gtgtggaaag
     4681 tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc
     4741 aggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat
     4801 tagtcagcaa ccatagtccc gcccctaact ccgcccatcc cgcccctaac tccgcccagt
     4861 tccgcccatt ctccgcccca tggctgacta atttttttta tttatgcaga ggccgaggcc
     4921 gcctctgcct ctgagctatt ccagaagtag tgaggaggct tttttggagg cctaggcttt
     4981 tgcaaaaagc tcccgggagc ttgtatatcc attttcggat ctgatcaaga gacaggatga
     5041 ggatcgtttc gcatgattga acaagatgga ttgcacgcag gttctccggc cgcttgggtg
     5101 gagaggctat tcggctatga ctgggcacaa cagacaatcg gctgctctga tgccgccgtg
     5161 ttccggctgt cagcgcaggg gcgcccggtt ctttttgtca agaccgacct gtccggtgcc
     5221 ctgaatgaac tgcaggacga ggcagcgcgg ctatcgtggc tggccacgac gggcgttcct
     5281 tgcgcagctg tgctcgacgt tgtcactgaa gcgggaaggg actggctgct attgggcgaa
     5341 gtgccggggc aggatctcct gtcatctcac cttgctcctg ccgagaaagt atccatcatg
     5401 gctgatgcaa tgcggcggct gcatacgctt gatccggcta cctgcccatt cgaccaccaa
     5461 gcgaaacatc gcatcgagcg agcacgtact cggatggaag ccggtcttgt cgatcaggat
     5521 gatctggacg aagagcatca ggggctcgcg ccagccgaac tgttcgccag gctcaaggcg
     5581 cgcatgcccg acggcgagga tctcgtcgtg acccatggcg atgcctgctt gccgaatatc
     5641 atggtggaaa atggccgctt ttctggattc atcgactgtg gccggctggg tgtggcggac
     5701 cgctatcagg acatagcgtt ggctacccgt gatattgctg aagagcttgg cggcgaatgg
     5761 gctgaccgct tcctcgtgct ttacggtatc gccgctcccg attcgcagcg catcgccttc
     5821 tatcgccttc ttgacgagtt cttctgagcg ggactctggg gttcgaaatg accgaccaag
     5881 cgacgcccaa cctgccatca cgagatttcg attccaccgc cgccttctat gaaaggttgg
     5941 gcttcggaat cgttttccgg gacgccggct ggatgatcct ccagcgcggg gatctcatgc
     6001 tggagttctt cgcccacccc aacttgttta ttgcagctta taatggttac aaataaagca
     6061 atagcatcac aaatttcaca aataaagcat ttttttcact gcattctagt tgtggtttgt
     6121 ccaaactcat caatgtatct tatcatgtct gtataccgtc gacctctagc tagagcttgg
     6181 cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta tccgctcaca attccacaca
     6241 acatacgagc cggaagcata aagtgtaaag cctggggtgc ctaatgagtg agctaactca
     6301 cattaattgc gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc
     6361 attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt
     6421 cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact
     6481 caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag
     6541 caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata
     6601 ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc
     6661 cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg
     6721 ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc
     6781 tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg
     6841 gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc
     6901 ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga
     6961 ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg
     7021 gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa
     7081 aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt ttttttgttt
     7141 gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta
     7201 cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat
     7261 caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa
     7321 gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct
     7381 cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta
     7441 cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct
     7501 caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg
     7561 gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa
     7621 gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt
     7681 cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta
     7741 catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca
     7801 gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta
     7861 ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct
     7921 gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg
     7981 cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac
     8041 tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact
     8101 gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa
     8161 atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt
     8221 ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat
     8281 gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg
     8341 acgtc
//
