LOCUS       sequence-337997-        7418 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasmid for endogenous C-terminal tagging of the Tetrahymena 
            thermophila Outer Dynein Arm intermediate chain 3 with FLAG and ZZ 
            tags..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7418)
  AUTHORS   Mali GR, Ali FA, Lau CK, Begum F, Boulanger J, Howe JD, Chen ZA, 
            Rappsilber J, Skehel M, Carter AP
  TITLE     Shulin packages axonemal outer dynein arms for ciliary targeting.
  JOURNAL   Science. 2021 Feb 26;371(6532):910-916. doi: 
            10.1126/science.abe0526.
  PUBMED    33632841
REFERENCE   2  (bases 1 to 7418)
  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: "10.1126/science"; 
            journalName: "Science"; date: "2021-02-26- 26"; volume: "371"; 
            issue: "6532"; pages: "910-916"
COMMENT     Sequence Label: pBluescript-IC3-FLAG-Tev-ZZ
FEATURES             Location/Qualifiers
     source          1..7418
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        21..125
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(126..194,195..986)
                     /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     complement(344..363)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1157..1745
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1646..1665
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1899..1916
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    2033..2054
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(2069..2074,2075..2092,2093..2099)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2107..2123
                     /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)."
     primer_bind     2112..2134
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     2131..2147
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2131..2147
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        2168..2186
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     CDS             3059..3124
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             3128..3148
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             4849..5643
                     /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="MASLDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4903..4922)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5513..5532
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(6774..6793)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(6775..6793)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(6800..6817)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(6800..6816)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6809..6831)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      6958..7413
                     /direction=RIGHT
                     /label=M13 ori
                     /note="M13 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(7045..7064)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     7255..7276
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
ORIGIN
        1 gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa
       61 atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga
      121 agagtatgag tattcaacat ttccgtgtcg cccttattcc cttttttgcg gcattttgcc
      181 ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg
      241 gtgcacgagt gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc
      301 gccccgaaga acgttttcca atgatgagca cttttaaagt tctgctatgt ggcgcggtat
      361 tatcccgtat tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg
      421 acttggttga gtactcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag
      481 aattatgcag tgctgccata accatgagtg ataacactgc ggccaactta cttctgacaa
      541 cgatcggagg accgaaggag ctaaccgctt ttttgcacaa catgggggat catgtaactc
      601 gccttgatcg ttgggaaccg gagctgaatg aagccatacc aaacgacgag cgtgacacca
      661 cgatgcctgt agcaatggca acaacgttgc gcaaactatt aactggcgaa ctacttactc
      721 tagcttcccg gcaacaatta atagactgga tggaggcgga taaagttgca ggaccacttc
      781 tgcgctcggc ccttccggct ggctggttta ttgctgataa atctggagcc ggtgagcgtg
      841 ggtctcgcgg tatcattgca gcactggggc cagatggtaa gccctcccgt atcgtagtta
      901 tctacacgac ggggagtcag gcaactatgg atgaacgaaa tagacagatc gctgagatag
      961 gtgcctcact gattaagcat tggtaactgt cagaccaagt ttactcatat atactttaga
     1021 ttgatttaaa acttcatttt taatttaaaa ggatctaggt gaagatcctt tttgataatc
     1081 tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa
     1141 agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa
     1201 aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc
     1261 cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt
     1321 agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc
     1381 tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac
     1441 gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca
     1501 gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg
     1561 ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag
     1621 gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt
     1681 ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat
     1741 ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc
     1801 acatgttctt tcctgcgtta tcccctgatt ctgtggataa ccgtattacc gcctttgagt
     1861 gagctgatac cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg agcgaggaag
     1921 cggaagagcg cccaatacgc aaaccgcctc tccccgcgcg ttggccgatt cattaatgca
     1981 gctggcacga caggtttccc gactggaaag cgggcagtga gcgcaacgca attaatgtga
     2041 gttagctcac tcattaggca ccccaggctt tacactttat gcttccggct cgtatgttgt
     2101 gtggaattgt gagcggataa caatttcaca caggaaacag ctatgaccat gattacgcca
     2161 agctcgaaat taaccctcac taaagggaac aaaagctgga gctccaccgc ggtggcggcc
     2221 gctctagagt ctgggattac tactacagac aaaacgaaat cgctttctct cacaaagtta
     2281 gtgactcacc tttaacatgc attaaaatta actaaactgg tggtgcctac cataattctg
     2341 gtaagctctg tgctattggt gattaagatg gtaaacaatc agataaattt tttattgttg
     2401 taagtattaa tttttattgt taaaaaaata ggtactgtta ctattcttga attatgtgat
     2461 tctctttaca caatgtaacc caaggaaaaa gatattatta acgaagtaaa tagttttgaa
     2521 ttaaaagttt aaataattaa aaaataatat aaataaatga tagatgttcg aaagagaata
     2581 cagaaaggaa aagaacttgg aaacaatcaa aaagtaatag gaattagcta aaagataagt
     2641 ttaaaaagat atgggtagcc aaaaagaaaa gtgggaaaag aagaaacttg aaatgattga
     2701 aactgctgaa gccagtttcc atgaaaatct tgctaagaac cctgttaatg aagaagaatt
     2761 taatgaactt gactcacctt ctgaaaagag aaagaaaact aattaaaatt aaggtagaga
     2821 ataagaagaa taaagcagag aagaataaga agcatctggt aacttcaatt aacaacaata
     2881 gtaataataa gaagaagaat agtagtaaga aggggaataa taacaccatt aaaatcaaga
     2941 acaccaaaac ggataaggac atgaaaatgg ctaagaggaa ggtgaagaaa atggtgaaga
     3001 aggtaattaa taagaaaatg aaggataaga agaaaacgaa taataataag aaggatccga
     3061 ttacaaagat catgatggtg attacaaaga tcacgatatt gattacaaag atgatgatga
     3121 taaagctgaa aatctttatt ttcaaggtgc ttctgctcaa catgatgaag ctgttgataa
     3181 caaatttaat aaggaacaac aaaatgcttt ttacgaaatt ttacatcttc ctaatttaaa
     3241 tgaagaacaa agaaatgctt tcattcaatc attaaaagat gatccttcac aatctgctaa
     3301 tttattagct gaagctaaaa aattaaatga tgctcaagct cctaaagttg ataataaatt
     3361 caacaaagaa caacaaaacg ctttttatga aattcttcat ttacctaacc ttaatgaaga
     3421 acaaagaaac gctttcattc aatctttaaa ggatgatcct tctcaatctg ctaatttact
     3481 tgctgaagct aaaaagttaa atggtgctca agctcctaag gttgatgcta atactagttg
     3541 agcgaactga atcggtcagc taaaccaacc aatcaacata ataaacttta ttatttttac
     3601 ttaagcatct tactgttgtt gtaatagtag agaaagaaat acccaattaa cttcattcac
     3661 ataacattaa tatctataaa catctttttt ctcacatata tacaactctc taaatcaaca
     3721 aataactttt taaaaataat ggatatatat taacaaaata atatatctct ttttacaaaa
     3781 tagttcttat ataaatacgt attctgcact cacccgcatt tttcacaaca aaaacatacc
     3841 aaaaaaattc ttacttctac atgtttcctt tcttattatt acaaaattat tttataaata
     3901 gcatacaaaa ataaatacaa taaaaaaata aacaaaatcc agatcccccg ggctgcagga
     3961 attcgatata caatttattt ctaaaaaata tttaaaaata aaaaataata agggttttga
     4021 ataactcctt taatttaaat acacattttt aaattttttt tagctcttta aatattcata
     4081 aaaataaaaa ataactaact aaaaataaat aaaaagataa taatgattaa aggtataata
     4141 ctgaataaga aaaaacataa tagagtactt attttttata tcactatttt taatatcttg
     4201 aaagcaaaac ttttttatat atcttaaaat atattgtatc gtttattcaa ttattttctt
     4261 taaatttcaa atatattgat aaaaaagatg acatgttttt taaagaaaac atgaaatata
     4321 aaatagataa atatcaatta ttttatttat taaatatata agctgctcaa aacatagctc
     4381 attcatcaat tataatatgt gaatcattaa ttttcaaaat attactcatt atttaggcta
     4441 tcatttattt tttattttca attatccgtt tctattatat tttaatatta agttgtgatt
     4501 cttgaatttt gtgtcatgaa ttatttgtaa atctttttat ttctgataaa aaatataaat
     4561 tgattgactc atgatttaaa tcatgagtca acctaactaa ttttcaaaat tcttctattc
     4621 taaaatatag atgtgattct tgaatctctc ttgaatataa agtaattttt tatatttctg
     4681 atataattct tagctacgtg attcacgatt tatgcaatga tccatataaa ataatgtaaa
     4741 tagtgtatat atatatattc gtctttttta ttctttatat aatttaaaaa aattaaaaaa
     4801 atttaataaa gctctaataa aataaataat aatactaaac ttaaaataat ggcaagcttg
     4861 gatggattgc acgcaggttc tccggccgct tgggtggaga ggctattcgg ctatgactgg
     4921 gcacaacaga caatcggctg ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc
     4981 ccggttcttt ttgtcaagac cgacctgtcc ggtgccctga atgaactgca ggacgaggca
     5041 gcgcggctat cgtggctggc cacgacgggc gttccttgcg cagctgtgct cgacgttgtc
     5101 actgaagcgg gaagggactg gctgctattg ggcgaagtgc cggggcagga tctcctgtca
     5161 tctcaccttg ctcctgccga gaaagtatcc atcatggctg atgcaatgcg gcggctgcat
     5221 acgcttgatc cggctacctg cccattcgac caccaagcga aacatcgcat cgagcgagca
     5281 cgtactcgga tggaagccgg tcttgtcgat caggatgatc tggacgaaga gcatcagggg
     5341 ctcgcgccag ccgaactgtt cgccaggctc aaggcgcgca tgcccgacgg cgaggatctc
     5401 gtcgtgaccc atggcgatgc ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct
     5461 ggattcatcg actgtggccg gctgggtgtg gcggaccgct atcaggacat agcgttggct
     5521 acccgtgata ttgctgaaga gcttggcggc gaatgggctg accgcttcct cgtgctttac
     5581 ggtatcgccg ctcccgattc gcagcgcatc gccttctatc gccttcttga cgagttcttc
     5641 tgagatcctt aaattaaaaa ttcaatatat atttacaaac tttcatataa aataaatata
     5701 ttatataaaa ttaattttta gtgtattata ttaacattaa agcaccaaaa aaacgtgtta
     5761 atatactact ataaaatata atttattcca aattgactaa aatcattatt ttacaactca
     5821 tttgtatata tattttatgt caattatttt ttttaacttt ctaaaaaaaa aaattcctct
     5881 tcacatacat gttagctctt aaaaatttgt ctgcaaatcc aataataata tttttttttg
     5941 ccattaaatt ttcaaatttt tactggaaaa atgcaagacc aatagatttt tgaaaaatta
     6001 ataaaaaatg tatactcttt cttgattata tacaataagt agatttttgt tttgttaact
     6061 gtatgattaa catacttaat attatataaa aaaactacac caaaatcaaa ttgcaaaatt
     6121 aattatattc acaatattta tgttcatatt actaaagttg tatatttata aaaaataaat
     6181 ttatttaaaa taaaataata tttttattat atttgcttat ttacatgagt gtgctcttac
     6241 aaacttaaat ctatattgaa ttaattggct aaataattat taaatttaaa taaaaatttt
     6301 atactaatta taaatttgtt gattttttga attagcataa aaattataaa tataaaaaga
     6361 aaagaaaaaa actcttctct atttattgtt ccttaatttt tactttactc tctaaagtta
     6421 tttataacta ataaaattat ttcataaata tttaaaattt aatctcttgt gaacataaat
     6481 tttagagaaa aaaaaaggtg aggtagaatt ataatcaata aaataataaa tatgtaattt
     6541 aattgaaata taaattaaaa taagtttatt agatttttac attctaaatt tgtagattgt
     6601 tatttattta ttttttctaa aaaataatat ttaattaaat taaccacttt gctacaaaat
     6661 ttacatttta acagtttata tttactttct aatgaaatta atgagagttg ttattagagt
     6721 ttatttattt aacaataatt aaattctcga gggggggccc ggtacccaat tcgccctata
     6781 gtgagtcgta ttacaattca ctggccgtcg ttttacaacg tcgtgactgg gaaaaccctg
     6841 gcgttaccca acttaatcgc cttgcagcac atcccccttt cgccagctgg cgtaatagcg
     6901 aagaggcccg caccgatcgc ccttcccaac agttgcgcag cctgaatggc gaatgggacg
     6961 cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta
     7021 cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt
     7081 tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg
     7141 ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt agtgggccat
     7201 cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac
     7261 tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt gatttataag
     7321 ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg
     7381 cgaattttaa caaaatatta acgcttacaa tttaggtg
//
