LOCUS       sequence-348158-        4345 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  This plasmid is used for generating C-terminal knock-in plasmid 
            and/or PCR donors in zebrafish..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4345)
  AUTHORS   Levic DS, Yamaguchi N, Wang S, Knaut H, Bagnat M
  TITLE     Knock-in tagging in zebrafish facilitated by insertion into 
            non-coding regions.
  JOURNAL   Development. 2021 Sep 8. pii: 272090. doi: 10.1242/dev.199994.
  PUBMED    34495314
REFERENCE   2  (bases 1 to 4345)
  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: 
            "Development. 2021 Sep 8. pii: 272090. doi: 10.1242/dev.199994."
COMMENT     Sequence Label: pUC19-TgKI-MCS-tdTomato-polyA
FEATURES             Location/Qualifiers
     source          1..4345
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    1486..1502
                     /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(1510..1516,1517..1534,1535..1540))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1555..1576
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1693..1710)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1864..2452)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1944..1963)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2623..3414,3415..3483))
                     /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     3246..3265
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3484..3588)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3656..3674
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(3712..3734)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     3834..3853
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4047..4069
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     4061..4078
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     4062..4078
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             join(4155..4345,1..1237)
                     /codon_start=1
                     /product="tandem dimeric (pseudo-monomeric) derivative of 
                     DsRed (Shaner et al., 2004)"
                     /label=tdTomato
                     /note="mammalian codon-optimized"
                     /translation="VSKGEEVIKEFMRFKVRMEGSMNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYKKLSFPEGFKWERVMNFEDGGL
                     VTVTQDSSLQDGTLIYKVKMRGTNFPPDGPVMQKKTMGWEASTERLYPRDGVLKGEIHQ
                     ALKLKDGGHYLVEFKTIYMAKKPVQLPGYYYVDTKLDITSHNEDYTIVEQYERSEGRHH
                     LFLGHGTGSTGSGSSGTASSEDNNMAVIKEFMRFKVRMEGSMNGHEFEIEGEGEGRPYE
                     GTQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYKKLSFPEGFKWERVMN
                     FEDGGLVTVTQDSSLQDGTLIYKVKMRGTNFPPDGPVMQKKTMGWEASTERLYPRDGVL
                     KGEIHQALKLKDGGHYLVEFKTIYMAKKPVQLPGYYYVDTKLDITSHNEDYTIVEQYER
                     SEGRHHLFLYGMDELYK"
ORIGIN
        1 gttcatgtac ggctccaagg cgtacgtgaa gcaccccgcc gacatccccg attacaagaa
       61 gctgtccttc cccgagggct tcaagtggga gcgcgtgatg aacttcgagg acggcggtct
      121 ggtgaccgtg acccaggact cctccctgca ggacggcacg ctgatctaca aggtgaagat
      181 gcgcggcacc aacttccccc ccgacggccc cgtaatgcag aagaagacca tgggctggga
      241 ggcctccacc gagcgcctgt acccccgcga cggcgtgctg aagggcgaga tccaccaggc
      301 cctgaagctg aaggacggcg gccactacct ggtggagttc aagaccatct acatggccaa
      361 gaagcccgtg caactgcccg gctactacta cgtggacacc aagctggaca tcacctccca
      421 caacgaggac tacaccatcg tggaacagta cgagcgctcc gagggccgcc accacctgtt
      481 cctggggcat ggcaccggca gcaccggcag cggcagctcc ggcaccgcct cctccgagga
      541 caacaacatg gccgtcatca aagagttcat gcgcttcaag gtgcgcatgg agggctccat
      601 gaacggccac gagttcgaga tcgagggcga gggcgagggc cgcccctacg agggcaccca
      661 gaccgccaag ctgaaggtga ccaagggcgg ccccctgccc ttcgcctggg acatcctgtc
      721 cccccagttc atgtacggct ccaaggcgta cgtgaagcac cccgccgaca tccccgatta
      781 caagaagctg tccttccccg agggcttcaa gtgggagcgc gtgatgaact tcgaggacgg
      841 cggtctggtg accgtgaccc aggactcctc cctgcaggac ggcacgctga tctacaaggt
      901 gaagatgcgc ggcaccaact tcccccccga cggccccgta atgcagaaga agaccatggg
      961 ctgggaggcc tccaccgagc gcctgtaccc ccgcgacggc gtgctgaagg gcgagatcca
     1021 ccaggccctg aagctgaagg acggcggcca ctacctggtg gagttcaaga ccatctacat
     1081 ggccaagaag cccgtgcaac tgcccggcta ctactacgtg gacaccaagc tggacatcac
     1141 ctcccacaac gaggactaca ccatcgtgga acagtacgag cgctccgagg gccgccacca
     1201 cctgttcctg tacggcatgg acgagctgta caagtaaatc attctcagta tcccctgctt
     1261 atgcactcac cactttttcc cattatttta ttcatgttgt gttgctaaat tcaactacat
     1321 tcgtattacc atgttgtttc atttgttttt tttttatctc agctgtcaaa ataaaagcat
     1381 tgaaatacaa cattgttcta tttttcattg tggttttgca tgcccggtaa ataaaaaaac
     1441 aaggtaataa aaataagatg cgaaatgaga agtatatctg tgaaattgtt atccgctcac
     1501 aattccacac aacatacgag ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt
     1561 gagctaactc acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc
     1621 gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg
     1681 ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt
     1741 atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa
     1801 gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc
     1861 gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag
     1921 gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt
     1981 gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg
     2041 aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg
     2101 ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg
     2161 taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac
     2221 tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg
     2281 gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt
     2341 taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg
     2401 tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc
     2461 tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt
     2521 ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt
     2581 taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag
     2641 tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt
     2701 cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg caatgatacc
     2761 gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc
     2821 cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg
     2881 ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac
     2941 aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg
     3001 atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc
     3061 tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact
     3121 gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc
     3181 aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat
     3241 acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc
     3301 ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac
     3361 tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa
     3421 aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact
     3481 catactcttc ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg
     3541 atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg
     3601 aaaagtgcca cctgacgtct aagaaaccat tattatcatg acattaacct ataaaaatag
     3661 gcgtatcacg aggccctttc gtctcgcgcg tttcggtgat gacggtgaaa acctctgaca
     3721 catgcagctc ccggagacgg tcacagcttg tctgtaagcg gatgccggga gcagacaagc
     3781 ccgtcagggc gcgtcagcgg gtgttggcgg gtgtcggggc tggcttaact atgcggcatc
     3841 agagcagatt gtactgagag tgcaccatat gcggtgtgaa ataccgcaca gatgcgtaag
     3901 gagaaaatac cgcatcaggc gccattcgcc attcaggctg cgcaactgtt gggaagggcg
     3961 atcggtgcgg gcctcttcgc tattacgcca gctggcgaaa gggggatgtg ctgcaaggcg
     4021 attaagttgg gtaacgccag ggttttccca gtcacgacgt tgtaaaacga cggccagtgg
     4081 cgagggcgat gccacctacg ggatgcggcc gcaccggtgg atccgaattc gctagcccgc
     4141 ggtctagact cgaggtgagc aagggcgagg aggtcatcaa agagttcatg cgcttcaagg
     4201 tgcgcatgga gggctccatg aacggccacg agttcgagat cgagggcgag ggcgagggcc
     4261 gcccctacga gggcacccag accgccaagc tgaaggtgac caagggcggc cccctgccct
     4321 tcgcctggga catcctgtcc cccca
//
