LOCUS       sequence-174537-        6886 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  TALEN-mediated mutagenesis of Exon 2 in NvFP-7R.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6886)
  AUTHORS   Ikmi A, McKinney SA, Delventhal KM, Gibson MC
  TITLE     TALEN and CRISPR/Cas9-mediated genome editing in the early-branching
            metazoan Nematostella vectensis.
  JOURNAL   Nat Commun. 2014 Nov 24;5:5486. doi: 10.1038/ncomms6486.
  PUBMED    25417547
REFERENCE   2  (bases 1 to 6886)
  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"; journalName: "Nat 
            Commun."; date: "2014-11-24"; volume: "5:5486. doi"; pages: " 
            10.1038/ncomms6486"
COMMENT     Sequence Label: pCS2TAL3_RR_NvFP-7R_Ex2-RIGHT
FEATURES             Location/Qualifiers
     source          1..6886
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        35..53
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     35..52
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     regulatory      85..94
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             97..117
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             118..207
                     /codon_start=1
                     /product="three tandem HA epitope tags"
                     /label=3xHA
                     /translation="YPYDVPDYAGYPYDVPDYAGSYPYDVPDYA"
     CDS             2311..2898
                     /codon_start=1
                     /product="nonspecific DNA cleavage domain of the FokI 
                     endonuclease (Li et al., 1992)"
                     /label=FokI cleavage domain
                     /note="must dimerize to be catalytically active"
                     /translation="QLVKSELEEKKSELRHKLKYVPHEYIELIEIARNSTQDRILEMKV
                     MEFFMKVYGYRGKHLGGSRKPDGAIYTVGSPIDYGVIVDTKAYSGGYNLPIGQAREMQR
                     YVEENQTRNKHINPNEWWKVYPSSVTEFKFLFVSGHFKGNYKAQLTRLNHITNCNGAVL
                     SVEELLIGGEMIKAGTLTLEEVRRKFNNGEINF"
     polyA_signal    2931..3065
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2955..2974)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3009..3028
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(3182..3202)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(3182..3200)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3221..3237)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3221..3237)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3234..3256)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3245..3261
                     /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(3269..3275,3276..3293,3294..3299))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3314..3335
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3452..3469)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3623..4211)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3703..3722)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4382..5173,5174..5242))
                     /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     5005..5024
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5243..5347)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(5373..5828)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5510..5531)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5722..5741
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        5902..6886
                     /label=CMV IE94 promoter
                     /note="enhancer/promoter region of simian cytomegalovirus 
                     major immediate early transcription unit IE94"
ORIGIN
        1 cgccattctg cctggggacg tcggagcaag cttgatttag gtgacactat agaatacaag
       61 ctacttgttc tttttgcagg atctgccacc atggctccaa agaagaagcg taaggtatac
      121 ccatacgatg ttcctgacta tgcgggctat ccctatgacg tcccggacta tgcaggatcg
      181 tatccatatg acgttccaga ttacgctgct catggtaccg tggatctacg cacgctcggc
      241 tacagtcagc agcagcaaga gaagatcaaa ccgaaggtgc gttcgacagt ggcgcagcac
      301 cacgaggcac tggtgggcca tgggtttaca cacgcgcaca tcgttgcgct cagccaacac
      361 ccggcagcgt tagggaccgt cgctgtcacg tatcagcaca taatcacggc gttgccagag
      421 gcgacacacg aagacatcgt tggcgtcggc aaacagtggt ccggcgcacg cgccctggag
      481 gccttgctca cggatgcggg ggagttgaga ggtccgccgt tacagttgga cacaggccaa
      541 cttgtgaaga ttgcaaaacg tggcggcgtg accgcaatgg aggcagtgca tgcatcgcgc
      601 aatgcactga cgggtgcccc cctgaacctg accccggacc aagtggtggc tatcgccagc
      661 aacaatggcg gcaagcaagc gctcgaaacg gtgcagcggc tgttgccggt gctgtgccag
      721 gaccatggcc tgaccccgga ccaagtggtg gctatcgcca gcaacattgg cggcaagcaa
      781 gcgctcgaaa cggtgcagcg gctgttgccg gtgctgtgcc aggaccatgg cctgactccg
      841 gaccaagtgg tggctatcgc cagccacgat ggcggcaagc aagcgctcga aacggtgcag
      901 cggctgttgc cggtgctgtg ccaggaccat ggcctgaccc cggaccaagt ggtggctatc
      961 gccagcaaca atggcggcaa gcaagcgctc gaaacggtgc agcggctgtt gccggtgctg
     1021 tgccaggacc atggcctgac cccggaccaa gtggtggcta tcgccagcaa caatggcggc
     1081 aagcaagcgc tcgaaacggt gcagcggctg ttgccggtgc tgtgccagga ccatggcctg
     1141 accccggacc aagtggtggc tatcgccagc aacattggcg gcaagcaagc gctcgaaacg
     1201 gtgcagcggc tgttgccggt gctgtgccag gaccatggcc tgactccgga ccaagtggtg
     1261 gctatcgcca gccacgatgg cggcaagcaa gcgctcgaaa cggtgcagcg gctgttgccg
     1321 gtgctgtgcc aggaccatgg cctgactccg gaccaagtgg tggctatcgc cagccacgat
     1381 ggcggcaagc aagcgctcga aacggtgcag cggctgttgc cggtgctgtg ccaggaccat
     1441 ggcctgactc cggaccaagt ggtggctatc gccagccacg atggcggcaa gcaagcgctc
     1501 gaaacggtgc agcggctgtt gccggtgctg tgccaggacc atggcctgac tccggaccaa
     1561 gtggtggcta tcgccagcca cgatggcggc aagcaagcgc tcgaaacggt gcagcggctg
     1621 ttgccggtgc tgtgccagga ccatggcctg accccggacc aagtggtggc tatcgccagc
     1681 aacggtggcg gcaagcaagc gctcgaaacg gtgcagcggc tgttgccggt gctgtgccag
     1741 gaccatggcc tgactccgga ccaagtggtg gctatcgcca gccacgatgg cggcaagcaa
     1801 gcgctcgaaa cggtgcagcg gctgttgccg gtgctgtgcc aggaccatgg cctgactccg
     1861 gaccaagtgg tggctatcgc cagccacgat ggcggcaagc aagcgctcga aacggtgcag
     1921 cggctgttgc cggtgctgtg ccaggaccat ggcctgaccc cggaccaagt ggtggctatc
     1981 gccagcaaca ttggcggcaa gcaagcgctc gaaacggtgc agcggctgtt gccggtgctg
     2041 tgccaggacc atggcctgac cccggaccaa gtggtggcta tcgccagcaa cggtggcggc
     2101 aagcaagcgc tcgaaagcat tgtggcccag ctgagccggc ctgatccggc gttggccgcg
     2161 ttgaccaacg accacctcgt cgccttggcc tgcctcggcg gacgtcctgc catggatgca
     2221 gtgaaaaagg gattgccgca cgcgccggaa ttgatcagaa gagtcaatcg ccgtattggc
     2281 gaacgcacgt cccatcgcgt tgccggatcc caactagtca aaagtgaact ggaggagaag
     2341 aaatctgaac ttcgtcataa attgaaatat gtgcctcatg aatatattga attaattgaa
     2401 attgccagaa attccactca ggatagaatt cttgaaatga aggtaatgga attttttatg
     2461 aaagtttatg gatatagagg taaacatttg ggtggatcaa ggaaaccgga cggagcaatt
     2521 tatactgtcg gatctcctat tgattacggt gtgatcgtgg atactaaagc ttatagcgga
     2581 ggttataatc tgccaattgg ccaagcacgt gaaatgcaac gatatgtcga agaaaatcaa
     2641 acacgaaaca aacatatcaa ccctaatgaa tggtggaaag tctatccatc ttctgtaacg
     2701 gaatttaagt ttttatttgt gagtggtcac tttaaaggaa actacaaagc tcagcttaca
     2761 cgattaaatc atatcactaa ttgtaatgga gctgttctta gtgtagaaga gcttttaatt
     2821 ggtggagaaa tgattaaagc cggcacatta accttagagg aagtgcgccg gaaatttaat
     2881 aacggcgaga taaactttta atctagaact atagtgagtc gtattacgta gatccagaca
     2941 tgataagata cattgatgag tttggacaaa ccacaactag aatgcagtga aaaaaatgct
     3001 ttatttgtga aatttgtgat gctattgctt tatttgtaac cattataagc tgcaataaac
     3061 aagttaacaa caacaattgc attcatttta tgtttcaggt tcagggggag gtgtgggagg
     3121 ttttttaatt cgcggccgcg gcgccaatgc attgggcccg gtacgtaccc agcttttgtt
     3181 ccctttagtg agggttaatt gcgcgcttgg cgtaatcatg gtcatagctg tttcctgtgt
     3241 gaaattgtta tccgctcaca attccacaca acatacgagc cggaagcata aagtgtaaag
     3301 cctggggtgc ctaatgagtg agctaactca cattaattgc gttgcgctca ctgcccgctt
     3361 tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag
     3421 gcggtttgcg tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg
     3481 ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat
     3541 caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta
     3601 aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa
     3661 atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc
     3721 cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt
     3781 ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca
     3841 gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg
     3901 accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat
     3961 cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta
     4021 cagagttctt gaagtggtgg cctaactacg gctacactag aaggacagta tttggtatct
     4081 gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac
     4141 aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa
     4201 aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa
     4261 actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt
     4321 taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca
     4381 gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca
     4441 tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc
     4501 ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa
     4561 accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc
     4621 agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca
     4681 acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat
     4741 tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag
     4801 cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac
     4861 tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt
     4921 ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt
     4981 gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc
     5041 tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat
     5101 ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca
     5161 gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga
     5221 cacggaaatg ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg
     5281 gttattgtct catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg
     5341 ttccgcgcac atttccccga aaagtgccac ctaaattgta agcgttaata ttttgttaaa
     5401 attcgcgtta aatttttgtt aaatcagctc attttttaac caataggccg aaatcggcaa
     5461 aatcccttat aaatcaaaag aatagaccga gatagggttg agtgttgttc cagtttggaa
     5521 caagagtcca ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa ccgtctatca
     5581 gggcgatggc ccactacgtg aaccatcacc ctaatcaagt tttttggggt cgaggtgccg
     5641 taaagcacta aatcggaacc ctaaagggag cccccgattt agagcttgac ggggaaagcc
     5701 ggcgaacgtg gcgagaaagg aagggaagaa agcgaaagga gcgggcgcta gggcgctggc
     5761 aagtgtagcg gtcacgctgc gcgtaaccac cacacccgcc gcgcttaatg cgccgctaca
     5821 gggcgcgtcc cattcgccat tcaggctgcg caactgttgg gaagggcgat cggtgcgggc
     5881 ctcttcgcta ttacgccagt cgaccatagc caattcaata tggcgtatat ggactcatgc
     5941 caattcaata tggtggatct ggacctgtgc caattcaata tggcgtatat ggactcgtgc
     6001 caattcaata tggtggatct ggaccccagc caattcaata tggcggactt ggcaccatgc
     6061 caattcaata tggcggactt ggcactgtgc caactgggga ggggtctact tggcacggtg
     6121 ccaagtttga ggaggggtct tggccctgtg ccaagtccgc catattgaat tggcatggtg
     6181 ccaataatgg cggccatatt ggctatatgc caggatcaat atataggcaa tatccaatat
     6241 ggccctatgc caatatggct attggccagg ttcaatacta tgtattggcc ctatgccata
     6301 tagtattcca tatatgggtt ttcctattga cgtagatagc ccctcccaat gggcggtccc
     6361 atataccata tatggggctt cctaataccg cccatagcca ctcccccatt gacgtcaatg
     6421 gtctctatat atggtctttc ctattgacgt catatgggcg gtcctattga cgtatatggc
     6481 gcctccccca ttgacgtcaa ttacggtaaa tggcccgcct ggctcaatgc ccattgacgt
     6541 caataggacc acccaccatt gacgtcaatg ggatggctca ttgcccattc atatccgttc
     6601 tcacgccccc tattgacgtc aatgacggta aatggcccac ttggcagtac atcaatatct
     6661 attaatagta acttggcaag tacattacta ttggaaggac gccagggtac attggcagta
     6721 ctcccattga cgtcaatggc ggtaaatggc ccgcgatggc tgccaagtac atccccattg
     6781 acgtcaatgg ggaggggcaa tgacgcaaat gggcgttcca ttgacgtaaa tgggcggtag
     6841 gcgtgcctaa tgggaggtct atataagcaa tgctcgttta gggaac
//
