LOCUS       sequence-111829-        6522 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  expresses FLAGHA-tagged HuD in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6522)
  TITLE     Identification of the transcriptome-wide mRNA targets of the 
            neuronal ELAVL proteins (unpublished)
REFERENCE   2  (bases 1 to 6522)
  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"
COMMENT     Sequence Label: pFRT-TODestFLAGHA_HuD
FEATURES             Location/Qualifiers
     source          1..6522
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             1..42
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     primer_bind     complement(86..105)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        277..656
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        657..860
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     811..831
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    862..880
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    883..901
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     911..935
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             1035..1058
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             1059..1085
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     protein_bind    1101..1125
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(2276..2300)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             2353..2394
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     primer_bind     complement(2474..2491)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2480..2704
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(2837..2856)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    2988..3035
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     CDS             3043..4065
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             3043..4065
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     polyA_signal    4195..4316
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4232..4251)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4286..4305
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4365..4381)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4365..4381)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4378..4400)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4389..4405
                     /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(4413..4419,4420..4437,4438..4443))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4458..4479
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4596..4613)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4767..5355)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4847..4866)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5526..6317,6318..6386))
                     /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     6149..6168
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6387..6491)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ggtaagccta tccctaaccc tctcctcggt ctcgattcta cggacggatc gggagatctc
       61 ccgatcccct atggtgcact ctcagtacaa tctgctctga tgccgcatag ttaagccagt
      121 atctgctccc tgcttgtgtg ttggaggtcg ctgagtagtg cgcgagcaaa atttaagcta
      181 caacaaggca aggcttgacc gacaattgca tgaagaatct gcttagggtt aggcgttttg
      241 cgctgcttcg cgatgtacgg gccagatata cgcgttgaca ttgattattg actagttatt
      301 aatagtaatc aattacgggg tcattagttc atagcccata tatggagttc cgcgttacat
      361 aacttacggt aaatggcccg cctggctgac cgcccaacga cccccgccca ttgacgtcaa
      421 taatgacgta tgttcccata gtaacgccaa tagggacttt ccattgacgt caatgggtgg
      481 agtatttacg gtaaactgcc cacttggcag tacatcaagt gtatcatatg ccaagtacgc
      541 cccctattga cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag tacatgacct
      601 tatgggactt tcctacttgg cagtacatct acgtattagt catcgctatt accatggtga
      661 tgcggttttg gcagtacatc aatgggcgtg gatagcggtt tgactcacgg ggatttccaa
      721 gtctccaccc cattgacgtc aatgggagtt tgttttggaa ccaaaatcaa cgggactttc
      781 caaaatgtcg taacaactcc gccccattga cgcaaatggg cggtaggcgt gtacggtggg
      841 aggtctatat aagcagagct ctccctatca gtgatagaga tctccctatc agtgatagag
      901 atcgtcgacg agctcgttta gtgaaccgtc agatcgcctg gagacgccat ccacgctgtt
      961 ttgacctcca tagaagacac cgggaccgat ccagcctccg gactctagcg tttaaactta
     1021 agcttggtac catggactac aaggacgacg atgacaagta cccttatgac gtgcccgatt
     1081 acgctttgaa ccagatatca acaagtttgt acaaaaaagc aggctccacc atgggaacca
     1141 attcagtcga catggttatg ataattagca ccatggagcc tcaggtgtca aatggtccga
     1201 catccaatac aagcaatgga ccctccagca acaacagaaa ctgtccttct cccatgcaaa
     1261 caggggcaac cacagatgac agcaaaacca acctcatcgt caactattta ccccagaata
     1321 tgacccaaga agaattcagg agtctcttcg ggagcattgg tgaaatagaa tcctgcaaac
     1381 ttgtgagaga caaaattaca ggacagagtt tagggtatgg atttgttaac tatattgatc
     1441 caaaggatgc agagaaagcc atcaacactt taaatggact cagactccag accaaaacca
     1501 taaaggtctc atatgcccgt ccgagctctg cctcaatcag ggatgctaac ctctatgtta
     1561 gcggccttcc caaaaccatg acccagaagg aactggagca acttttctcg caatacggcc
     1621 gtatcatcac ctcacgaatc ctggttgatc aagtcacagg agtgtccaga ggggtgggat
     1681 tcatccgctt tgataagagg attgaggcag aagaagccat caaagggctg aatggccaga
     1741 agcccagcgg tgctacggaa ccgattactg tgaagtttgc caacaacccc agccagaagt
     1801 ccagccaggc cctgctctcc cagctctacc agtcccccaa ccggcgctac ccaggtccac
     1861 ttcaccacca ggctcagagg ttcaggctgg acaatttgct taatatggcc tatggcgtaa
     1921 agaggttctc cccaattacc attgatggaa tgacaagcct tgtgggaatg aacatccctg
     1981 gtcacacagg aactgggtgg tgcatctttg tctacaacct gtcccccgat tccgatgaga
     2041 gtgtcctctg gcagctcttt ggcccctttg gagcagtgaa caacgtaaag gtgattcgtg
     2101 acttcaacac caacaagtgc aagggattcg gctttgtcac catgaccaac tatgatgagg
     2161 cggccatggc catcgccagc ctcaacgggt accgcctggg agacagagtg ttgcaagttt
     2221 cctttaaaac caacaaagcc cacaagtcct gagcggccgc actcgagata tctagaccca
     2281 gctttcttgt acaaagtggt tgatatccag cacagtggcg gccgctcaag tctagagggc
     2341 ccgcggttcg aaggtaagcc tatccctaac cctctcctcg gtctcgattc tacgcgtacc
     2401 tgatccgcgg ccgcatagat aactgatcca gtgtgcctcg agtctagagg gcccgtttaa
     2461 acccgctgat cagcctcgac tgtgccttct agttgccagc catctgttgt ttgcccctcc
     2521 cccgtgcctt ccttgaccct ggaaggtgcc actcccactg tcctttccta ataaaatgag
     2581 gaaattgcat cgcattgtct gagtaggtgt cattctattc tggggggtgg ggtggggcag
     2641 gacagcaagg gggaggattg ggaagacaat agcaggcatg ctggggatgc ggtgggctct
     2701 atggcttctg aggcggaaag aaccagctgg ggctctaggg ggtatcccca cgcgccctgt
     2761 agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc
     2821 agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc
     2881 tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg
     2941 cacctcgacc ccaaaaaact tgattagggt gatggttcac gtacctagaa gttcctattc
     3001 cgaagttcct attctctaga aagtatagga acttccttgg ccaaaaagcc tgaactcacc
     3061 gcgacgtctg tcgagaagtt tctgatcgaa aagttcgaca gcgtctccga cctgatgcag
     3121 ctctcggagg gcgaagaatc tcgtgctttc agcttcgatg taggagggcg tggatatgtc
     3181 ctgcgggtaa atagctgcgc cgatggtttc tacaaagatc gttatgttta tcggcacttt
     3241 gcatcggccg cgctcccgat tccggaagtg cttgacattg gggaattcag cgagagcctg
     3301 acctattgca tctcccgccg tgcacagggt gtcacgttgc aagacctgcc tgaaaccgaa
     3361 ctgcccgctg ttctgcagcc ggtcgcggag gccatggatg cgatcgctgc ggccgatctt
     3421 agccagacga gcgggttcgg cccattcgga ccgcaaggaa tcggtcaata cactacatgg
     3481 cgtgatttca tatgcgcgat tgctgatccc catgtgtatc actggcaaac tgtgatggac
     3541 gacaccgtca gtgcgtccgt cgcgcaggct ctcgatgagc tgatgctttg ggccgaggac
     3601 tgccccgaag tccggcacct cgtgcacgcg gatttcggct ccaacaatgt cctgacggac
     3661 aatggccgca taacagcggt cattgactgg agcgaggcga tgttcgggga ttcccaatac
     3721 gaggtcgcca acatcttctt ctggaggccg tggttggctt gtatggagca gcagacgcgc
     3781 tacttcgagc ggaggcatcc ggagcttgca ggatcgccgc ggctccgggc gtatatgctc
     3841 cgcattggtc ttgaccaact ctatcagagc ttggttgacg gcaatttcga tgatgcagct
     3901 tgggcgcagg gtcgatgcga cgcaatcgtc cgatccggag ccgggactgt cgggcgtaca
     3961 caaatcgccc gcagaagcgc ggccgtctgg accgatggct gtgtagaagt actcgccgat
     4021 agtggaaacc gacgccccag cactcgtccg agggcaaagg aatagcacgt actacgagat
     4081 ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc
     4141 ggctggatga tcctccagcg cggggatctc atgctggagt tcttcgccca ccccaacttg
     4201 tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa
     4261 gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttatcat
     4321 gtctgtatac cgtcgacctc tagctagagc ttggcgtaat catggtcata gctgtttcct
     4381 gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag cataaagtgt
     4441 aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc
     4501 gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg
     4561 agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg
     4621 gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca
     4681 gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac
     4741 cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac
     4801 aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg
     4861 tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac
     4921 ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat
     4981 ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag
     5041 cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac
     5101 ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt
     5161 gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaaggac agtatttggt
     5221 atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc
     5281 aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga
     5341 aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac
     5401 gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc
     5461 cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct
     5521 gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca
     5581 tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct
     5641 ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca
     5701 ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc
     5761 atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg
     5821 cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct
     5881 tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa
     5941 aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta
     6001 tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc
     6061 ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg
     6121 agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa
     6181 gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg
     6241 agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc
     6301 accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg
     6361 gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat
     6421 cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata
     6481 ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg tc
//
