LOCUS       sequence-183796-        7058 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  TALEN Backbone for Phaeodactylum tricornutum_ fcpA promotor_NN.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7058)
  AUTHORS   Serif M, Lepetit B, Weissert K, Kroth PG, Rio Bartulos C
  TITLE     A fast and reliable strategy to generate TALEN-mediated gene 
            knockouts in the diatom Phaeodactylum tricornutum
  JOURNAL   Algal Research Volume 23, April 2017, Pages 186–195
REFERENCE   2  (bases 1 to 7058)
  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: "Algal 
            Research Volume 23, April 2017, Pages 186–195"
COMMENT     Sequence Label: pM9_fcpA_NN
FEATURES             Location/Qualifiers
     source          1..7058
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        528..546
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      548..557
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             557..622
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             629..649
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     promoter        join(1118..1123,1124..1141,1142..1148)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     CDS             1202..1861
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(1268..1287)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     CDS             2203..2508
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     2387..2406
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             2812..3399
                     /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
                     MEFFMKVYGYRGKHLGGSRKPDGAIYTVGSPIDYGVIVDTKAYSGGYNLPIGQADEMQR
                     YVEENQTRNKHINPNEWWKVYPSSVTEFKFLFVSGHFKGNYKAQLTRLNHITNCNGAVL
                     SVEELLIGGEMIKAGTLTLEEVRRKFNNGEINF"
     primer_bind     complement(3417..3434)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     CDS             4068..4442
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     promoter        complement(4694..4712)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4799..4816)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4970..5558)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5050..5069)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5729..6520,6521..6589))
                     /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     6352..6371
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6590..6694)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6762..6780
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(6818..6840)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     6940..6959
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        7040..7058
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     7040..7057
                     /label=SP6
                     /note="SP6 promoter, forward primer"
ORIGIN
        1 accagatccc ccgggctgca ggacgcaatg gaggattatc accgcaaaaa tgaacttcga
       61 aaaaaacttt cgagcgacca tggaaaagga ggatcagatt cagattacaa cagtggattg
      121 ctctggtagc aaatatcttc tgctagattg gctcatggtc ggttttggac gttcgaagct
      181 caccgtcaaa agaaacaaaa gagaagaatg acgtcttcgt gacgtagaat ctacgactgt
      241 actcggatct gggaaatgaa ttgactcacg gtcttcttcg agtcctgtta caggcccttg
      301 gtccgaaccc ccacacgatt tttgcaccaa agatttgctt caatttgctg gatgttttga
      361 ctgcaagatc agctggccta gcaagagtgc tcgtgttgct tcgtcgggaa tccctacgaa
      421 tttcagttct gcacaaattt gtctgccgtt tcgagaattc gatatcatcg actaattcga
      481 gctctctggc taactagaga acccactgct tactggctta tcgaaattaa tacgactcac
      541 tataggggcc accatggact ataaggacca cgacggagac tacaaggatc atgatattga
      601 ttacaaagac gatgacgata agatggcccc aaagaagaag cggaaggtcg gtatccacgg
      661 agtcccagca gccgtagatt tgagaacttt gggatattca cagcagcagc aggaaaagat
      721 caagcccaaa gtgaggtcga cagtcgcgca gcatcacgaa gcgctggtgg gtcatgggtt
      781 tacacatgcc cacatcgtag ccttgtcgca gcaccctgca gcccttggca cggtcgccgt
      841 caagtaccag gacatgattg cggcgttgcc ggaagccaca catgaggcga tcgtcggtgt
      901 ggggaaacag tggagcggag cccgagcgct tgaggccctg ttgacggtcg cgggagagct
      961 gagagggcct ccccttcagc tggacacggg ccagttgctg aagatcgcga agcggggagg
     1021 agtcacggcg gtcgaggcgg tgcacgcgtg gcgcaatgcg ctcacgggag cacccctcaa
     1081 cctgacagag accgcggccg cattaggcac cccaggcttt acactttatg cttccggctc
     1141 gtataatgtg tggattttga gttaggatcc gtcgagattt tcaggagcta aggaagctaa
     1201 aatggagaaa aaaatcactg gatataccac cgttgatata tcccaatggc atcgtaaaga
     1261 acattttgag gcatttcagt cagttgctca atgtacctat aaccagaccg ttcagctgga
     1321 tattacggcc tttttaaaga ccgtaaagaa aaataagcac aagttttatc cggcctttat
     1381 tcacattctt gcccgcctga tgaatgctca tccggaattc cgtatggcaa tgaaagacgg
     1441 tgagctggtg atatgggata gtgttcaccc ttgttacacc gttttccatg agcaaactga
     1501 aacgttttca tcgctctgga gtgaatacca cgacgatttc cggcagtttc tacacatata
     1561 ttcgcaagat gtggcgtgtt acggtgaaaa cctggcctat ttccctaaag ggtttattga
     1621 gaatatgttt ttcgtctcag ccaatccctg ggtgagtttc accagttttg atttaaacgt
     1681 ggccaatatg gacaacttct tcgcccccgt tttcaccatg ggcaaatatt atacgcaagg
     1741 cgacaaggtg ctgatgccgc tggcgattca ggttcatcat gccgtttgtg atggcttcca
     1801 tgtcggcaga atgcttaatg aattacaaca gtactgcgat gagtggcagg gcggggcgta
     1861 aagatctgga tccggcttac taaaagccag ataacagtat gcgtatttgc gcgctgattt
     1921 ttgcggtata agaatatata ctgatatgta tacccgaagt atgtcaaaaa gaggtatgct
     1981 atgaagcagc gtattacagt gacagttgac agcgacagct atcagttgct caaggcatat
     2041 atgatgtcaa tatctccggt ctggtaagca caaccatgca gaatgaagcc cgtcgtctgc
     2101 gtgccgaacg ctggaaagcg gaaaatcagg aagggatggc tgaggtcgcc cggtttattg
     2161 aaatgaacgg ctcttttgct gacgagaaca ggggctggtg aaatgcagtt taaggtttac
     2221 acctataaaa gagagagccg ttatcgtctg tttgtggatg tacagagtga tattattgac
     2281 acgcccgggc gacggatggt gatccccctg gccagtgcac gtctgctgtc agataaagtc
     2341 tcccgtgaac tttacccggt ggtgcatatc ggggatgaaa gctggcgcat gatgaccacc
     2401 gatatggcca gtgtgccggt ctccgttatc ggggaagaag tggctgatct cagccaccgc
     2461 gaaaatgaca tcaaaaacgc cattaacctg atgttctggg gaatataaat gtcaggctcc
     2521 cttatacaca gccagtctgc aggtcgacgg tctcgactca cgcctgagca ggtagtggct
     2581 attgcatcca acaacggggg cagacccgca ctggagtcaa tcgtggccca gctttcgagg
     2641 ccggaccccg cgctggccgc actcactaat gatcatcttg tagcgctggc ctgcctcggc
     2701 ggacgacccg ccttggatgc ggtgaagaag gggctcccgc acgcgcctgc attgattaag
     2761 cggaccaaca gaaggattcc cgagaggaca tcacatcgag tggcaggttc ccaactcgtg
     2821 aagagtgaac ttgaggagaa aaagtcggag ctgcggcaca aattgaaata cgtaccgcat
     2881 gaatacatcg aacttatcga aattgctagg aactcgactc aagacagaat ccttgagatg
     2941 aaggtaatgg agttctttat gaaggtttat ggataccgag ggaagcatct cggtggatca
     3001 cgaaaacccg acggagcaat ctatacggtg gggagcccga ttgattacgg agtgatcgtc
     3061 gacacgaaag cctacagcgg tgggtacaat cttcccatcg ggcaggcaga tgagatgcaa
     3121 cgttatgtcg aagaaaatca gaccaggaac aaacacatca atccaaatga gtggtggaaa
     3181 gtgtatcctt catcagtgac cgagtttaag tttttgtttg tctctgggca tttcaaaggc
     3241 aactataagg cccagctcac acggttgaat cacattacga actgcaatgg tgcggttttg
     3301 tccgtagagg aactgctcat tggtggagaa atgatcaaag cgggaactct gacactggaa
     3361 gaagtcagac gcaagtttaa caatggcgag atcaatttcc gctcataaaa aatcagcctc
     3421 gactgtgcct tctagttgcc agccatctgt tgtttgcccc tcccccgtgc cttccttgac
     3481 cctggaagct tcagaagcgt gctatcgaac tcaaccaggg acgtgcggca caaatgggca
     3541 tccttgctct catggtgcac gaacagttgg gagtctctat ccttccttaa aaatttaatt
     3601 ttcattagtt gcagtcactc cgctttggtt tcacagtcag gaataacact agctcgtctt
     3661 caccatggat gccaatctcg cctattcatg gtgtataaaa gttcaacatc caaagctaga
     3721 acttttggaa agagaaagaa tatccgaata gggcacggcg tgccgtattg ttggagtgga
     3781 ctagcagaaa gtgaggaagg cacaggatga gttttctcga gacatacctt cagcgtcgtc
     3841 ttcactgtca cagtcaactg acagtaatcg ttgatccgga gagattcaaa attcaatctg
     3901 tttggacctg gataagacac aagagcgaca tcctgacatg aacgccgtaa acagcaaatc
     3961 ctggttgaac acgtatcctt ttgggggcct cccgctacga cgctcgctcc agctggggct
     4021 tccttactat acacagcgcg catatttcac ggttgccaga tgtcaagatg gccaagttga
     4081 ccagtgccgt tccggtgctc accgcgcgcg acgtcgccgg agcggtcgag ttctggaccg
     4141 accggctcgg gttctcccgg gacttcgtgg aggacgactt cgccggtgtg gtccgggacg
     4201 acgtgaccct gttcatcagc gcggtccagg accaggtggt gccggacaac accctggcct
     4261 gggtgtgggt gcgcggcctg gacgagctgt acgccgagtg gtcggaggtc gtgtccacga
     4321 acttccggga cgcctccggg ccggccatga ccgagatcgg cgagcagccg tgggggcggg
     4381 agttcgccct gcgcgacccg gccggcaact gcgtgcactt cgtggccgag gagcaggact
     4441 gaaccttcct taaaaattta attttcatta gttgcagtca ctccgctttg gtttcacagt
     4501 caggaataac actagctcgt cttcaccatg gatgccaatc tcgcctattc atggtgtata
     4561 aaagttcaac atccaaagct agaacttttg gaaagagaaa gaatatccga atagggcacg
     4621 gcgtgccgta ttgttggagt ggactagcag aaagtgagga aggcacagga tgagttttct
     4681 cgaggccgga ctccctatag tgagtcgtat taatttcgat aagccaggtt aacctgcatt
     4741 aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct tccgcttcct
     4801 cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa
     4861 aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa
     4921 aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc
     4981 tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga
     5041 caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc
     5101 cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt
     5161 ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct
     5221 gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg
     5281 agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta
     5341 gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct
     5401 acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa
     5461 gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt
     5521 gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta
     5581 cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat
     5641 caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa
     5701 gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct
     5761 cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta
     5821 cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcgg gacccacgct
     5881 caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg
     5941 gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa
     6001 gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt
     6061 cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta
     6121 catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca
     6181 gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta
     6241 ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct
     6301 gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg
     6361 cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac
     6421 tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact
     6481 gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa
     6541 atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt
     6601 ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat
     6661 gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg
     6721 acgtctaaga aaccattatt atcatgacat taacctataa aaataggcgt atcacgaggc
     6781 cctttcgtct cgcgcgtttc ggtgatgacg gtgaaaacct ctgacacatg cagctcccgg
     6841 agacggtcac agcttgtctg taagcggatg ccgggagcag acaagcccgt cagggcgcgt
     6901 cagcgggtgt tggcgggtgt cggggctggc ttaactatgc ggcatcagag cagattgtac
     6961 tgagagtgca ccatatggac atattgtcgt tagaacgcgg ctacaattaa tacataacct
     7021 tatgtatcat acacatacga tttaggtgac actataga
//
