LOCUS       Exported                6525 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  expresses FLAGHA-tagged HuC in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    pFRT-TODestFLAGHA_HuC
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6525)
  TITLE     Identification of the transcriptome-wide mRNA targets of the 
            neuronal ELAVL proteins (unpublished)
REFERENCE   2  (bases 1 to 6525)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6525
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     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(2279..2303)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             2356..2397
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     primer_bind     complement(2477..2494)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2483..2707
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(2840..2859)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    2991..3038
                     /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             3046..4068
                     /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    4198..4319
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4235..4254)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4289..4308
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4368..4384)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4368..4384)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4381..4403)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4392..4408
                     /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(4416..4446)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4461..4482
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4599..4616)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4770..5358)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4850..4869)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(5529..6389)
                     /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     6152..6171
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6390..6494)
                     /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 catggtcact cagatactgg gggccatgga gtctcaggtg ggggggggcc
     1201 cggccggccc ggccctgccc aacgggccac tccttggtac aaatggagcc actgacgaca
     1261 gcaagaccaa cctcatcgtc aactacctgc cccagaacat gacccaggat gagttcaaga
     1321 gtctcttcgg cagcattggc gacatcgagt cctgcaagtt ggttcgggac aagatcacag
     1381 ggcagagcct tggctacggg tttgtgaact attctgaccc caatgatgca gacaaagcca
     1441 tcaacaccct caacggcctc aaattacaga cgaagaccat caaggtgtcc tatgccagac
     1501 ccagttcagc atccatccgg gatgctaacc tgtacgtcag cgggctcccc aagaccatga
     1561 gccagaaaga gatggagcag ctcttctccc agtacggccg catcatcacg tcccgcatcc
     1621 tggtggacca ggtcacaggt gtctctcggg gtgtgggatt catccgcttt gacaagagga
     1681 ttgaggccga agaggctatc aaaggactga atgggcagaa gccgctgggc gcagctgagc
     1741 ccatcacagt caagttcgcg aacaacccaa gtcagaagac ggggcaggcg ctgctcaccc
     1801 acctctacca gtcatccgcc cggcgctacg caggccccct acaccatcag acccagcgtt
     1861 tccggctgga caatttgctc aacatggcct acggcgtcaa gagtcccctg tcgctcatcg
     1921 ccaggttctc gccgatcgcc atcgatggta tgagcggcct ggcgggcgtg ggcctgtcgg
     1981 ggggcgcggc gggcgccggc tggtgcatct tcgtgtacaa cctgtcaccg gaggcagacg
     2041 agagcgtgct gtggcagctg ttcgggcctt ttggggcagt caccaacgtc aaggtcatcc
     2101 gtgatttcac caccaacaag tgcaagggtt tcggcttcgt gaccatgacc aactatgacg
     2161 aggcggccat ggccatcgcc agcctgaacg gctatcgcct gggcgagcgc gtgctgcagg
     2221 tctccttcaa gaccagcaaa cagcacaagg cgtgagcggc cgcactcgag atatctagac
     2281 ccagctttct tgtacaaagt ggttgatatc cagcacagtg gcggccgctc aagtctagag
     2341 ggcccgcggt tcgaaggtaa gcctatccct aaccctctcc tcggtctcga ttctacgcgt
     2401 acctgatccg cggccgcata gataactgat ccagtgtgcc tcgagtctag agggcccgtt
     2461 taaacccgct gatcagcctc gactgtgcct tctagttgcc agccatctgt tgtttgcccc
     2521 tcccccgtgc cttccttgac cctggaaggt gccactccca ctgtcctttc ctaataaaat
     2581 gaggaaattg catcgcattg tctgagtagg tgtcattcta ttctgggggg tggggtgggg
     2641 caggacagca agggggagga ttgggaagac aatagcaggc atgctgggga tgcggtgggc
     2701 tctatggctt ctgaggcgga aagaaccagc tggggctcta gggggtatcc ccacgcgccc
     2761 tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt
     2821 gccagcgccc tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc
     2881 ggctttcccc gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta
     2941 cggcacctcg accccaaaaa acttgattag ggtgatggtt cacgtaccta gaagttccta
     3001 ttccgaagtt cctattctct agaaagtata ggaacttcct tggccaaaaa gcctgaactc
     3061 accgcgacgt ctgtcgagaa gtttctgatc gaaaagttcg acagcgtctc cgacctgatg
     3121 cagctctcgg agggcgaaga atctcgtgct ttcagcttcg atgtaggagg gcgtggatat
     3181 gtcctgcggg taaatagctg cgccgatggt ttctacaaag atcgttatgt ttatcggcac
     3241 tttgcatcgg ccgcgctccc gattccggaa gtgcttgaca ttggggaatt cagcgagagc
     3301 ctgacctatt gcatctcccg ccgtgcacag ggtgtcacgt tgcaagacct gcctgaaacc
     3361 gaactgcccg ctgttctgca gccggtcgcg gaggccatgg atgcgatcgc tgcggccgat
     3421 cttagccaga cgagcgggtt cggcccattc ggaccgcaag gaatcggtca atacactaca
     3481 tggcgtgatt tcatatgcgc gattgctgat ccccatgtgt atcactggca aactgtgatg
     3541 gacgacaccg tcagtgcgtc cgtcgcgcag gctctcgatg agctgatgct ttgggccgag
     3601 gactgccccg aagtccggca cctcgtgcac gcggatttcg gctccaacaa tgtcctgacg
     3661 gacaatggcc gcataacagc ggtcattgac tggagcgagg cgatgttcgg ggattcccaa
     3721 tacgaggtcg ccaacatctt cttctggagg ccgtggttgg cttgtatgga gcagcagacg
     3781 cgctacttcg agcggaggca tccggagctt gcaggatcgc cgcggctccg ggcgtatatg
     3841 ctccgcattg gtcttgacca actctatcag agcttggttg acggcaattt cgatgatgca
     3901 gcttgggcgc agggtcgatg cgacgcaatc gtccgatccg gagccgggac tgtcgggcgt
     3961 acacaaatcg cccgcagaag cgcggccgtc tggaccgatg gctgtgtaga agtactcgcc
     4021 gatagtggaa accgacgccc cagcactcgt ccgagggcaa aggaatagca cgtactacga
     4081 gatttcgatt ccaccgccgc cttctatgaa aggttgggct tcggaatcgt tttccgggac
     4141 gccggctgga tgatcctcca gcgcggggat ctcatgctgg agttcttcgc ccaccccaac
     4201 ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat
     4261 aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttat
     4321 catgtctgta taccgtcgac ctctagctag agcttggcgt aatcatggtc atagctgttt
     4381 cctgtgtgaa attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag
     4441 tgtaaagcct ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg
     4501 cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg
     4561 gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc
     4621 tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc
     4681 acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg
     4741 aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat
     4801 cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag
     4861 gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga
     4921 tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg
     4981 tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt
     5041 cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac
     5101 gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc
     5161 ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag gacagtattt
     5221 ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc
     5281 ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc
     5341 agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg
     5401 aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag
     5461 atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg
     5521 tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt
     5581 tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca
     5641 tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca
     5701 gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc
     5761 tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt
     5821 ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg
     5881 gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc
     5941 aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg
     6001 ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga
     6061 tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga
     6121 ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta
     6181 aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg
     6241 ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact
     6301 ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata
     6361 agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt
     6421 tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa
     6481 ataggggttc cgcgcacatt tccccgaaaa gtgccacctg acgtc
//
