LOCUS       sequence-73763-e        8549 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Right TALEN for PRKAG2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8549)
  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 8549)
  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-PRKAG2 Right TALEN
FEATURES             Location/Qualifiers
     source          1..8549
                     /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(4143..4160)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4149..4373
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      4419..4847
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4506..4525)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4716..4737
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4856..4876)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4861..5190
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5041..5176
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     5103..5122
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             5257..6051
                     /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(5311..5330)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5921..5940
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    6225..6346
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6262..6281)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6316..6335
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(6395..6411)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6395..6411)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6408..6430)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6419..6435
                     /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(6443..6449,6450..6467,6468..6473))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6488..6509
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6626..6643)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6797..7382)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6877..6896)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7553..8344,8345..8413))
                     /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     8176..8195
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8414..8518)
                     /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 attggcggca agcaagcgct cgaaacggtg
     1501 cagcggctgt tgccggtgct gtgccaggac catggcctga ctccggacca agtggtggct
     1561 atcgccagcc acgatggcgg caagcaagcg ctcgaaacgg tgcagcggct gttgccggtg
     1621 ctgtgccagg accatggcct gactccggac caagtggtgg ctatcgccag ccacgatggc
     1681 ggcaagcaag cgctcgaaac ggtgcagcgg ctgttgccgg tgctgtgcca ggaccatggc
     1741 ctgactccgg accaagtggt ggctatcgcc agccacgatg gcggcaagca agcgctcgaa
     1801 acggtgcagc ggctgttgcc ggtgctgtgc caggaccatg gcctgacccc ggaccaagtg
     1861 gtggctatcg ccagcaacat tggcggcaag caagcgctcg aaacggtgca gcggctgttg
     1921 ccggtgctgt gccaggacca tggcctgact ccggaccaag tggtggctat cgccagccac
     1981 gatggcggca agcaagcgct cgaaacggtg cagcggctgt tgccggtgct gtgccaggac
     2041 catggcctga ccccggacca agtggtggct atcgccagca acattggcgg caagcaagcg
     2101 ctcgaaacgg tgcagcggct gttgccggtg ctgtgccagg accatggcct gaccccggac
     2161 caagtggtgg ctatcgccag caacattggc ggcaagcaag cgctcgaaac ggtgcagcgg
     2221 ctgttgccgg tgctgtgcca ggaccatggc ctgaccccgg accaagtggt ggctatcgcc
     2281 agcaacggtg gcggcaagca agcgctcgaa acggtgcagc ggctgttgcc ggtgctgtgc
     2341 caggaccatg gcctgacccc ggaccaagtg gtggctatcg ccagcaacat tggcggcaag
     2401 caagcgctcg aaacggtgca gcggctgttg ccggtgctgt gccaggacca tggcctgacc
     2461 ccggaccaag tggtggctat cgccagccac gatggcggca agcaagcgct cgaaacggtg
     2521 cagcggctgt tgccggtgct gtgccaggac catggcctga ccccggacca agtggtggct
     2581 atcgccagca acggtggcgg caagcaagcg ctcgaaacgg tgcagcggct gttgccggtg
     2641 ctgtgccagg accatggcct gaccccggac caagtggtgg ctatcgccag caacattggc
     2701 ggcaagcaag cgctcgaaac ggtgcagcgg ctgttgccgg tgctgtgcca ggaccatggc
     2761 ctgaccccgg accaagtggt ggctatcgcc agcaacggtg gcggcaagca agcgctcgaa
     2821 acggtgcagc ggctgttgcc ggtgctgtgc caggaccatg gcctgactcc ggaccaagtg
     2881 gtggctatcg ccagccacga tggcggcaag caagcgctcg aaacggtgca gcggctgttg
     2941 ccggtgctgt gccaggacca tggcctgacc ccggaccaag tggtggctat cgccagcaac
     3001 ggtggcggca agcaagcgct cgaaacggtg cagcggctgt tgccggtgct gtgccaggac
     3061 catggcctga ccccggacca agtggtggct atcgccagca acaatggcgg caagcaagcg
     3121 ctcgaaacgg tgcagcggct gttgccggtg ctgtgccagg accatggcct gactccggac
     3181 caagtggtgg ctatcgccag ccacgatggc ggcaagcaag cgctcgaaac ggtgcagcgg
     3241 ctgttgccgg tgctgtgcca ggaccatggc ctgaccccgg accaagtggt ggctatcgcc
     3301 agcaacggtg gcggcaagca agcgctcgaa agcattgtgg cccagctgag ccggcctgat
     3361 ccggcgttgg ccgcgttgac caacgaccat ctggtggcgt tggcatgtct tggtggacga
     3421 cccgcgctcg atgcagtcaa aaagggtctg cctcatgctc ccgcattgat caaaagaacc
     3481 aaccggcgga ttcccgagag aacttcccat cgagtcgcgg gatcccagct ggtgaagagc
     3541 gagctggagg agaagaagtc cgagctgcgg cacaagctga agtacgtgcc ccacgagtac
     3601 atcgagctga tcgagatcgc caggaacagc acccaggacc gcatcctgga gatgaaggtg
     3661 atggagttct tcatgaaggt gtacggctac aggggaaagc acctgggcgg aagcagaaag
     3721 cctgacggcg ccatctatac agtgggcagc cccatcgatt acggcgtgat cgtggacaca
     3781 aaggcctaca gcggcggcta caatctgcct atcggccagg ccgacgagat gcagagatac
     3841 gtggaggaga accagacccg gaataagcac atcaacccca acgagtggtg gaaggtgtac
     3901 cctagcagcg tgaccgagtt caagttcctg ttcgtgagcg gccacttcaa gggcaactac
     3961 aaggcccagc tgaccaggct gaaccacatc accaaccgca atggcgccgt gctgagcgtg
     4021 gaggagctgc tgatcggcgg cgagatgatc aaagccggca ccctgacact ggaggaggtg
     4081 cggcgcaagt tcaacaacgg cgagatcaac ttctgataac ttaagtttaa accgctgatc
     4141 agcctcgact gtgccttcta gttgccagcc atctgttgtt tgcccctccc ccgtgccttc
     4201 cttgaccctg gaaggtgcca ctcccactgt cctttcctaa taaaatgagg aaattgcatc
     4261 gcattgtctg agtaggtgtc attctattct ggggggtggg gtggggcagg acagcaaggg
     4321 ggaggattgg gaagacaata gcaggcatgc tggggatgcg gtgggctcta tggcttctga
     4381 ggcggaaaga accagctggg gctctagggg gtatccccac gcgccctgta gcggcgcatt
     4441 aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca gcgccctagc
     4501 gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct ttccccgtca
     4561 agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc acctcgaccc
     4621 caaaaaactt gattagggtg atggttcacg tagtgggcca tcgccctgat agacggtttt
     4681 tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc aaactggaac
     4741 aacactcaac cctatctcgg tctattcttt tgatttataa gggattttgc cgatttcggc
     4801 ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaattaat tctgtggaat
     4861 gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc
     4921 atgcatctca attagtcagc aaccaggtgt ggaaagtccc caggctcccc agcaggcaga
     4981 agtatgcaaa gcatgcatct caattagtca gcaaccatag tcccgcccct aactccgccc
     5041 atcccgcccc taactccgcc cagttccgcc cattctccgc cccatggctg actaattttt
     5101 tttatttatg cagaggccga ggccgcctct gcctctgagc tattccagaa gtagtgagga
     5161 ggcttttttg gaggcctagg cttttgcaaa aagctcccgg gagcttgtat atccattttc
     5221 ggatctgatc aagagacagg atgaggatcg tttcgcatga ttgaacaaga tggattgcac
     5281 gcaggttctc cggccgcttg ggtggagagg ctattcggct atgactgggc acaacagaca
     5341 atcggctgct ctgatgccgc cgtgttccgg ctgtcagcgc aggggcgccc ggttcttttt
     5401 gtcaagaccg acctgtccgg tgccctgaat gaactgcagg acgaggcagc gcggctatcg
     5461 tggctggcca cgacgggcgt tccttgcgca gctgtgctcg acgttgtcac tgaagcggga
     5521 agggactggc tgctattggg cgaagtgccg gggcaggatc tcctgtcatc tcaccttgct
     5581 cctgccgaga aagtatccat catggctgat gcaatgcggc ggctgcatac gcttgatccg
     5641 gctacctgcc cattcgacca ccaagcgaaa catcgcatcg agcgagcacg tactcggatg
     5701 gaagccggtc ttgtcgatca ggatgatctg gacgaagagc atcaggggct cgcgccagcc
     5761 gaactgttcg ccaggctcaa ggcgcgcatg cccgacggcg aggatctcgt cgtgacccat
     5821 ggcgatgcct gcttgccgaa tatcatggtg gaaaatggcc gcttttctgg attcatcgac
     5881 tgtggccggc tgggtgtggc ggaccgctat caggacatag cgttggctac ccgtgatatt
     5941 gctgaagagc ttggcggcga atgggctgac cgcttcctcg tgctttacgg tatcgccgct
     6001 cccgattcgc agcgcatcgc cttctatcgc cttcttgacg agttcttctg agcgggactc
     6061 tggggttcga aatgaccgac caagcgacgc ccaacctgcc atcacgagat ttcgattcca
     6121 ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga
     6181 tcctccagcg cggggatctc atgctggagt tcttcgccca ccccaacttg tttattgcag
     6241 cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt
     6301 cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttatcat gtctgtatac
     6361 cgtcgacctc tagctagagc ttggcgtaat catggtcata gctgtttcct gtgtgaaatt
     6421 gttatccgct cacaattcca cacaacatac gagccggaag cataaagtgt aaagcctggg
     6481 gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc gctttccagt
     6541 cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt
     6601 tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc
     6661 tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg
     6721 ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg
     6781 ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac
     6841 gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg
     6901 gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct
     6961 ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg
     7021 tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct
     7081 gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac
     7141 tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt
     7201 tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc
     7261 tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca
     7321 ccgctggtag cggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc
     7381 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt
     7441 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa
     7501 aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat
     7561 gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct
     7621 gactccccgt cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg
     7681 caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag
     7741 ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta
     7801 attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg
     7861 ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg
     7921 gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct
     7981 ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta
     8041 tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg
     8101 gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc
     8161 cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg
     8221 gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga
     8281 tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg
     8341 ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat
     8401 gttgaatact catactcttc ctttttcaat attattgaag catttatcag ggttattgtc
     8461 tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca
     8521 catttccccg aaaagtgcca cctgacgtc
//
