LOCUS       sequence-355655-        5219 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses zebrafish ythdf2 with 3x flag tag, for mRNA injection.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5219)
  AUTHORS   Kontur C, Jeong M, Cifuentes D, Giraldez AJ
  TITLE     Ythdf m(6)A Readers Function Redundantly during Zebrafish 
            Development.
  JOURNAL   Cell Rep. 2020 Dec 29;33(13):108598. doi: 
            10.1016/j.celrep.2020.108598.
  PUBMED    33378672
REFERENCE   2  (bases 1 to 5219)
  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: "Cell Rep.";
            date: "2020-12-29"; volume: "33(13):108598. doi"; pages: " 
            10.1016/j.celrep.2020.108598"
COMMENT     Sequence Label: pSP64T-ythdf2-3xflag
FEATURES             Location/Qualifiers
     source          1..5219
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(11..28)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(182..770)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(262..281)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(941..1732,1733..1801))
                     /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     1564..1583
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1802..1906)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     1974..1992
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(2030..2052)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     2152..2171
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2385..2404
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        2694..2712
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     2694..2711
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     5'UTR           2724..2766
                     /label=Xenopus globin 5'-UTR
                     /note="translational enhancer from the 5'-UTR of the major 
                     beta-globin gene of Xenopus laevis"
     regulatory      2777..2786
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2786..2851
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     3'UTR           4746..4871
                     /label=Xenopus globin 3'-UTR
                     /note="3'-UTR of the major beta-globin gene of Xenopus 
                     laevis"
     primer_bind     complement(4999..5015)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4999..5015)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5012..5034)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5023..5039
                     /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(5047..5053,5054..5071,5072..5077))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5092..5113
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
ORIGIN
        1 cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat
       61 cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga
      121 acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt
      181 ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt
      241 ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc
      301 gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa
      361 gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct
      421 ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta
      481 actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg
      541 gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc
      601 ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta
      661 ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg
      721 gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt
      781 tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg
      841 tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta
      901 aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg
      961 aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg
     1021 tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc
     1081 gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg
     1141 agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg
     1201 aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag
     1261 gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat
     1321 caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc
     1381 cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc
     1441 ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa
     1501 ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac
     1561 gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt
     1621 cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc
     1681 gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa
     1741 caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca
     1801 tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat
     1861 acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa
     1921 aagtgccacc tgacgtctaa gaaaccatta ttatcatgac attaacctat aaaaataggc
     1981 gtatcacgag gccctttcgt ctcgcgcgtt tcggtgatga cggtgaaaac ctctgacaca
     2041 tgcagctccc ggagacggtc acagcttgtc tgtaagcgga tgccgggagc agacaagccc
     2101 gtcagggcgc gtcagcgggt gttggcgggt gtcggggctg gcttaactat gcggcatcag
     2161 agcagattgt actgagagtg caccattcga cgctctccct tatgcgactc ctgcattagg
     2221 aagcagccca gtagtaggtt gaggccgttg agcaccgccg ccgcaaggaa tggtgcatgc
     2281 aaggagatgg cgcccaacag tcccccggcc acggggcctg ccaccatacc cacgccgaaa
     2341 caagcgctca tgagcccgaa gtggcgagcc cgatcttccc catcggtgat gtcggcgata
     2401 taggcgccag caaccgcacc tgtggcgccg gtgatgccgg ccacgatgcg tccggcgtag
     2461 aggatctggc tagcgatgac cctgctgatt ggttcgctga ccatttccgg gtgcgggacg
     2521 gcgttaccag aaactcagaa ggttcgtcca accaaaccga ctctgacggc agtttacgag
     2581 agagatgata gggtctgctt cagtaagcca gatgctacac aattaggctt gtacatattg
     2641 tcgttagaac gcggctacaa ttaatacata accttatgta tcatacacat acgatttagg
     2701 tgacactata gaatacaagc ttgcttgttc tttttgcaga agctcagaat aaacgctcaa
     2761 ctttggcaga tccgcggccg ccatggacta caaagaccat gacggtgatt ataaagatca
     2821 tgacatcgac tacaaggatg acgatgacaa gaattcaggt tctggctcag gatcgtcagc
     2881 cagcagcctt ctggagcaga gaccgaaagg ccaagcaaac aaagtacaaa acggagctgt
     2941 tacccaaaag gatactttga acgatgatga gttcgagcct tacctgaatg ctcagcccag
     3001 acagagcaat gcctatacgg ccatgtccga ctcctacatg cccagttact acagcccctc
     3061 aataggcttc acctattccc tgaatgaagc ggcatggtca acaggtggag atcccccgat
     3121 gccttacctg gcctcttatg gacagcttag caatggggag caccactttc tcccggatgc
     3181 catgtttgga cagtctggag cactggggaa caaccctttc ctaggacagc atggcttcaa
     3241 cttctttccc agtgggattg actttcccgc ttggggcaac agcagctctc agggacagtc
     3301 cacacagagt tcaggttaca gcagcagtta cgcctatgcc cctagcacgc taggaggtgc
     3361 aatgattgac ggacagtctc cctttgcagc caatgagcca ctcaacaagg cagtggggat
     3421 gaatagcttg gatcagggca tggcagggct taaaattggg gcaggggaca tggcaccaaa
     3481 ggttgttggt tctggacttc ccggagggcc attgagtcag gtgtctacag ctccaaccat
     3541 gcctcctgcc tccatggctc ctgcgaaaac tgcttcttgg gcggacattg ccagcaagcc
     3601 agccaaacca cagcctaaat tgaaaactaa ggggggactt ggtgggacaa atctgccacc
     3661 acctccaata aaacataaca tggatattgg gacatgggac aacaagggga atatgcctaa
     3721 accagcagcc ccacagcaga cttccctgcc cactaatgga cagccaccca atcagtcgtc
     3781 tcctcaacca ggtgctactg ctggtggggt gcctcagttg cccctcagca atggacagct
     3841 agttcctcct actggacagc ttgtccagca ccctcttcct ccaggtggcc agcccggtgc
     3901 tgttccacct caactctccc agggcccacc agcttcccaa ccctcccagc caacccgctg
     3961 ggtgcccccg cgtaaccgtg ccaacggctt tggggatgca gcaggtggac ctggccaatc
     4021 ccctcccaat tcaggaatgg gtggcattac tgtgccagca gaaccccacc cagttctaga
     4081 aaaactgcgc atggtaaaca actacaatcc caaggacttc gactggaacc ctaaacatgg
     4141 tcgtgtcttc atcatcaaga gttactctga ggacgatatc caccggtcca ttaagtacaa
     4201 tatctggtgc agcacagaac atggcaataa gcgtctggac gctgcatacc gttcgctagc
     4261 caacaaaggg ccactgtacc tgctcttcag tgtgaatggc agtgggcact tctgtggcgt
     4321 ggcggagatg cgctcgcctg tggactacaa cacttgtgca ggcgtgtggt cgcaggacaa
     4381 gtggaagggt cgtttcgatg tgcgctggat ctttgtcaag gacgttccca acagccaact
     4441 aaggcacatt cgcttggaga acaatgagaa taagccggta accaactctc gcgacacaca
     4501 ggaggttcct ctagacaagg cacgtcaagt gttgaagatc atcgcgagtt acaagcacac
     4561 cacctccatt tttgacgact tctcacacta cgagaaacgt caggaagagg aggagagtgt
     4621 aaaaaaggtg gaggttcagg gcagtgaccc gtactccaat aactccagtc ggagccacta
     4681 cagaatgcag gatcgccaag gacgtgtaaa gtaactcgag aataattcgg atctggttac
     4741 cactaaacca gcctcaagaa cacccgaatg gagtctctaa gctacataat accaacttac
     4801 actttacaaa atgttgtccc ccaaaatgta gccattcgta tctgctccta ataaaaagaa
     4861 agtttcttca cattctaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa
     4921 aaaaaaammc cccccccccc ccccctgcag gtcgactcta gaggatcccc gggcgagctc
     4981 gaattaattc gtaatcatgt catagctgtt tcctgtgtga aattgttatc cgctcacaat
     5041 tccacacaac atacgagccg gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag
     5101 ctaactcaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg
     5161 ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgct
//
