LOCUS       sequence-68899-e        7009 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Golden Gate Compatible TALE Transcription Factor using VP64 Domain.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7009)
  TITLE     Bao Lab Golden Gate plasmids
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 7009)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: pTALE64
FEATURES             Location/Qualifiers
     source          1..7009
                     /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"
     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"
     CDS             complement(1159..1734)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITPSAQLTLTKGNKSWRTSRGGPVPNSPYSESYYARSLAVVL
                     QRRDWENPGVTQLNRLAAHPPFASWRNSEEARTDRPSQQLRSLNGEWDAPCSGARLQGG
                     TGERIAPGVHCLYRCYSPSLRDLQQLPGVELERRPPKLTRHSQQRLKCSRSRPLFTDPY
                     NCLVRGFGQGRNHILVFDSHRPKRRRVL"
     primer_bind     1584..1606
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     1598..1615
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     1599..1615
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        1625..1643
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1691..1711)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(1691..1709)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(1730..1746)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    1754..1770
                     /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(1778..1784,1785..1802,1803..1808))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1823..1844
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     CDS             2375..2395
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2420..2569
                     /codon_start=1
                     /product="tetrameric repeat of the minimal activation 
                     domain of herpes simplex virus VP16 (Beerli et al., 1998)"
                     /label=VP64
                     /translation="DALDDFDLDMLGSDALDDFDLDMLGSDALDDFDLDMLGSDALDDF
                     DLDML"
     primer_bind     complement(2603..2620)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2609..2833
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2879..3307
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2966..2985)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3176..3197
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3316..3336)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3321..3650
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3501..3636
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3563..3582
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3717..4511
                     /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(3771..3790)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4381..4400
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4685..4806
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4722..4741)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4776..4795
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4855..4871)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    4879..4895
                     /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(4903..4909,4910..4927,4928..4933))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4948..4969
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5086..5103)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5257..5842)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5337..5356)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6013..6804,6805..6873))
                     /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     6636..6655
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6874..6978)
                     /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 ggagacgggc
     1441 gccgctacag ggcgcgtccc attcgccatt caggctgcgc aactgttggg aagggcgatc
     1501 ggtgcgggcc tcttcgctat tacgccagct ggcgaaaggg ggatgtgctg caaggcgatt
     1561 aagttgggta acgccagggt tttcccagtc acgacgttgt aaaacgacgg ccagtgagcg
     1621 cgcgtaatac gactcactat agggcgaatt gggtaccggg ccccccctcg aggtcctcca
     1681 gcttttgttc cctttagtga gggttaattg cgcgcttggc gtaatcatgg tcatagctgt
     1741 ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa catacgagcc ggaagcataa
     1801 agtgtaaagc ctggggtgcc taatgagtga gctaactcac attaattgcg ttgcgctcac
     1861 tgcccgcttt ccaccggtcg tctccaacga ccatctggtg gcgttggcat gtcttggtgg
     1921 acgacccgcg ctcgatgcag tcaaaaaggg tctgcctcat gctcccgcat tgatcaaaag
     1981 aaccaaccgg cggattcccg agagaacttc ccatcgagtc gcggatcacg cgcaagtggt
     2041 ccgcgtgctc ggattcttcc agtgtcactc ccaccccgca caagcgttcg atgacgccat
     2101 gactcaattt ggtatgtcga gacacggact gctgcagctc tttcgtagag tcggtgtcac
     2161 agaactcgag gcccgctcgg gcacactgcc tcccgcctcc cagcggtggg acaggattct
     2221 ccaagcgagc ggtatgaaac gcgcgaagcc ttcacctacg tcaactcaga cacctgacca
     2281 ggcgagcctt catgcgttcg cagactcgct ggagagggat ttggacgcgc cctcgcccat
     2341 gcatgaaggg gaccaaactc gcgcgtcagc tagccccaag aagaagagaa aggtggaggc
     2401 cagcggttcc ggacgggctg acgcattgga cgattttgat ctggatatgc tgggaagtga
     2461 cgccctcgat gattttgacc ttgacatgct tggttcggat gcccttgatg actttgacct
     2521 cgacatgctc ggcagtgacg cccttgatga tttcgacctg gacatgctga ttaactctag
     2581 ttaagtttaa accgctgatc agcctcgact gtgccttcta gttgccagcc atctgttgtt
     2641 tgcccctccc ccgtgccttc cttgaccctg gaaggtgcca ctcccactgt cctttcctaa
     2701 taaaatgagg aaattgcatc gcattgtctg agtaggtgtc attctattct ggggggtggg
     2761 gtggggcagg acagcaaggg ggaggattgg gaagacaata gcaggcatgc tggggatgcg
     2821 gtgggctcta tggcttctga ggcggaaaga accagctggg gctctagggg gtatccccac
     2881 gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct
     2941 acacttgcca gcgccctagc gcccgctcct ttcgctttct tcccttcctt tctcgccacg
     3001 ttcgccggct ttccccgtca agctctaaat cgggggctcc ctttagggtt ccgatttagt
     3061 gctttacggc acctcgaccc caaaaaactt gattagggtg atggttcacg tagtgggcca
     3121 tcgccctgat agacggtttt tcgccctttg acgttggagt ccacgttctt taatagtgga
     3181 ctcttgttcc aaactggaac aacactcaac cctatctcgg tctattcttt tgatttataa
     3241 gggattttgc cgatttcggc ctattggtta aaaaatgagc tgatttaaca aaaatttaac
     3301 gcgaattaat tctgtggaat gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag
     3361 caggcagaag tatgcaaagc atgcatctca attagtcagc aaccaggtgt ggaaagtccc
     3421 caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccatag
     3481 tcccgcccct aactccgccc atcccgcccc taactccgcc cagttccgcc cattctccgc
     3541 cccatggctg actaattttt tttatttatg cagaggccga ggccgcctct gcctctgagc
     3601 tattccagaa gtagtgagga ggcttttttg gaggcctagg cttttgcaaa aagctcccgg
     3661 gagcttgtat atccattttc ggatctgatc aagagacagg atgaggatcg tttcgcatga
     3721 ttgaacaaga tggattgcac gcaggttctc cggccgcttg ggtggagagg ctattcggct
     3781 atgactgggc acaacagaca atcggctgct ctgatgccgc cgtgttccgg ctgtcagcgc
     3841 aggggcgccc ggttcttttt gtcaagaccg acctgtccgg tgccctgaat gaactgcagg
     3901 acgaggcagc gcggctatcg tggctggcca cgacgggcgt tccttgcgca gctgtgctcg
     3961 acgttgtcac tgaagcggga agggactggc tgctattggg cgaagtgccg gggcaggatc
     4021 tcctgtcatc tcaccttgct cctgccgaga aagtatccat catggctgat gcaatgcggc
     4081 ggctgcatac gcttgatccg gctacctgcc cattcgacca ccaagcgaaa catcgcatcg
     4141 agcgagcacg tactcggatg gaagccggtc ttgtcgatca ggatgatctg gacgaagagc
     4201 atcaggggct cgcgccagcc gaactgttcg ccaggctcaa ggcgcgcatg cccgacggcg
     4261 aggatctcgt cgtgacccat ggcgatgcct gcttgccgaa tatcatggtg gaaaatggcc
     4321 gcttttctgg attcatcgac tgtggccggc tgggtgtggc ggaccgctat caggacatag
     4381 cgttggctac ccgtgatatt gctgaagagc ttggcggcga atgggctgac cgcttcctcg
     4441 tgctttacgg tatcgccgct cccgattcgc agcgcatcgc cttctatcgc cttcttgacg
     4501 agttcttctg agcgggactc tggggttcga aatgaccgac caagcgacgc ccaacctgcc
     4561 atcacgagat ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt
     4621 ccgggacgcc ggctggatga tcctccagcg cggggatctc atgctggagt tcttcgccca
     4681 ccccaacttg tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt
     4741 cacaaataaa gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt
     4801 atcttatcat gtctgtatac cgtcgacctc tagctagagc ttggcgtaat catggtcata
     4861 gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag
     4921 cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg
     4981 ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca
     5041 acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc
     5101 gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg
     5161 gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa
     5221 ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga
     5281 cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag
     5341 ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct
     5401 taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg
     5461 ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc
     5521 ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt
     5581 aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta
     5641 tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac
     5701 agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc
     5761 ttgatccggc aaacaaacca ccgctggtag cggttttttt gtttgcaagc agcagattac
     5821 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca
     5881 gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac
     5941 ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac
     6001 ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt
     6061 tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt
     6121 accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt
     6181 atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc
     6241 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa
     6301 tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg
     6361 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt
     6421 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc
     6481 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt
     6541 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg
     6601 gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac
     6661 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc
     6721 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt
     6781 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg
     6841 aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat attattgaag
     6901 catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa
     6961 acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtc
//
