LOCUS       EW215_PGK-ZNF35(5-8)-de 5768 bp ds-DNA     circular     07-MAR-2025
DEFINITION  .
COMMENT     Concatenation of 4 sequences.
FEATURES             Location/Qualifiers
     misc_feature    8..521
                     /label="PGK promoter"
     misc_feature    127..127
                     /label="change to A?"
     primer          446..462
                     /label="PGK2"
                     /note="sequence: CATTCTGCACGCTTCAA"
     primer          515..529
                     /label="EWo203"
                     /note="sequence: cctgcagcccaagct"
     primer          515..540
                     /label="EWo209"
                     /note="sequence: cctgcagcccaagcttcgccaccatg"
     misc_feature    530..537
                     /label="Kozak"
     misc_feature    538..540
                     /label="start codon"
     CDS             538..540
                     /label="Translation 538-540"
                     /translation="M"
     misc_feature    541..861
                     /label="ZNF35 ZF Domains 5-8"
     CDS             541..861
                     /label="Translation 541-861"
                     /translation="YECNECGKTFTRSSNLIVHQRIHTGEKPFACNDCGKAFTQSANLIVHQRSHTGEKPYECKECGKAFSCFSHLIVHQRIHTAEKPYDCSECGKAFSQLSCLIVHQRIH"
     primer          complement(856..870)
                     /label="EWo196.1"
                     /note="sequence: tgcagcagcGTGAAT"
     primer          856..911
                     /label="EWo196.2"
                     /note="sequence: ATTCACgctgctgcaTGGGAGCTAggaATGtctggcggatctGAGGGCCAGAGCGA"
     CDS             862..897
                     /label="Translation 862-897"
                     /translation="AAAWELGMSGGS"
     misc_feature    898..1045
                     /label="N (NCOA3) Activation Domain"
     misc_feature    898..1353
                     /label="NZF Domain"
     CDS             898..1353
                     /label="Translation 898-1353"
                     /translation="EGQSDERALLDQLHTLLSNTDATGLEEIDRALGIPELVNQGQALEPKQDGGWGSMAEEFVTLKDVGMDFTLGDWEQLGLEQGDTFWDTALDNCQDLFLLDPPGGGDGLGLGHEKFPSDLDLDMFNGSLECDMESIIRSELMDADGLDFNFDS"
     primer          complement(1001..1015)
                     /label="EW_NZF_R"
                     /note="sequence: GGTTCACCAGCTCGG"
     misc_feature    1060..1203
                     /label="Z (ZNF473) Activation Domain"
     misc_feature    1228..1353
                     /label="F (FOXO3) Activation Domain"
     CDS             1354..1356
                     /label="Translation 1354-1356"
                     /translation="*"
     primer          complement(1359..1380)
                     /label="EKbR"
                     /note="sequence: ttgttcGGGCCctcctcgagcg"
     CDS             1375..1407
                     /label="c-myc epitope"
     misc_feature    1407..1409
                     /label="New Stop"
     CDS             1420..1437
                     /label="Polyhistidine tag"
     misc_feature    1438..1440
                     /label="STOP (missing now)"
     primer          complement(1441..1459)
                     /label="ToFlpR"
                     /note="sequence: CTGATCAGCGGGTTTAAAC"
     polyA_signal    1459..1673
                     /label="BGH polyadenylation sequence"
     primer          complement(1460..1477)
                     /label="BGHR Genewiz"
                     /note="sequence: TAGAAGGCACAGTCGAGG"
     primer          complement(1539..1558)
                     /label="BghR1"
                     /note="sequence: attaggaaaggacagtggga"
     primer          1650..1668
                     /label="bghF"
                     /note="sequence: AGACAATAGCAGGCATGCT"
     polyA_signal    1706..1729
                     /label="BGH pA"
     primer          complement(1808..1823)
                     /label="FlpInR"
                     /note="sequence: cgcgtaaccaccacac"
     primer          1941..1959
                     /label="FrtF"
                     /note="sequence: agggttccgatttagtgct"
     misc_feature    2013..2060
                     /label="FRT"
     gene            2068..3088
                     /label="Hygromycin"
     primer          3016..3034
                     /label="Hyg_F"
                     /note="sequence: ACCGATGGCTGTGTAGAAG"
     polyA_signal    3220..3350
                     /label="SV40 pA"
     primer          complement(3233..3258)
                     /label="SV40PA_R"
                     /note="sequence: GCTTTATTTGTAACCATTATAAGCTG"
     primer          complement(3257..3276)
                     /label="SV40pA-R Genewiz"
                     /note="sequence: GAAATTTGTGATGCTATTGC"
     primer          complement(3390..3406)
                     /label="M13R Genewiz"
                     /note="sequence: CAGGAAACAGCTATGAC"
     primer          complement(3404..3423)
                     /label="M13-48REV Genewiz"
                     /note="sequence: CGGATAACAATTTCACACAG"
     primer          3682..3701
                     /label="pUCori5F"
                     /note="sequence: AAGGCGGTAATACGGTTATC"
     rep_origin      3733..4406
                     /label="pUC ori"
     primer          complement(3853..3872)
                     /label="pUCoriR"
                     /note="sequence: tatagtcctgtcgggtttcg"
     CDS             complement(4551..5411)
                     /label="Amp (bla)"
     promoter        complement(5412..5510)
                     /label="bla promoter"
     primer          5623..5643
                     /label="FlpInF1"
                     /note="sequence: agtatctgctccctgcttgtg"
     primer          5687..5704
                     /label="FlpInF"
                     /note="sequence: aacaaggcaaggcttgac"
ORIGIN
        1 tacgcgttct accgggtagg ggaggcgctt ttcccaaggc agtctggagc atgcgcttta
       61 gcagccccgc tgggcacttg gcgctacaca agtggcctct ggcctcgcac acattccaca
      121 tccaccggta ggcgccaacc ggctccgttc tttggtggcc ccttcgcgcc accttctact
      181 cctcccctag tcaggaagtt cccccccgcc ccgcagctcg cgtcgtgcag gacgtgacaa
      241 atggaagtag cacgtctcat tagtctcgtg cagatggaca gcaccgctga gcaatggaag
      301 cgggtaggcc tttggggcag cggccaatag cagctttgct ccttcgcttt ctgggctcag
      361 aggctgggaa ggggtgggtc cgggggcggg ctcaggggcg ggctcagggg cggggcgggc
      421 gcccgaaggt cctccggagg cccggcattc tgcacgcttc aaaagcgcac gtctgccgcg
      481 ctgttctcct cttcctcatc tccgggcctt tcgacctgca gcccaagctt cgccaccatg
      541 TACGAGTGTA ATGAGTGCGG TAAGACCTTC ACACGATCTT CTAACCTTAT TGTGCATCAA
      601 CGAATCCATA CAGGCGAGAA GCCCTTTGCC TGCAATGACT GTGGGAAGGC TTTTACTCAG
      661 AGTGCCAACT TGATCGTGCA TCAGAGAAGC CATACGGGTG AGAAGCCCTA TGAATGCAAG
      721 GAATGCGGAA AGGCCTTCTC CTGCTTCAGC CACCTCATTG TACACCAGCG CATACACACG
      781 GCTGAGAAGC CATACGATTG CTCTGAATGT GGAAAGGCCT TTAGTCAGCT GTCTTGCTTG
      841 ATTGTGCATC AGAGAATTCA Cgctgctgca TGGGAGCTAg gaATGtctgg cggatctGAG
      901 GGCCAGAGCG ACGAGAGAGC CCTGCTGGAC CAGCTGCACA CCCTGCTGAG CAACACCGAC
      961 GCCACCGGCC TGGAGGAGAT CGACAGAGCC CTGGGCATCC CCGAGCTGGT GAACCAGGGC
     1021 CAGGCCCTGG AGCCCAAGCA GGACGGAGGC TGGGGATCTA TGGCCGAGGA ATTCGTGACC
     1081 CTGAAGGACG TGGGCATGGA CTTCACCCTG GGCGACTGGG AGCAGCTGGG CCTGGAGCAG
     1141 GGCGACACCT TCTGGGACAC CGCCCTGGAC AACTGCCAGG ACCTGTTCCT GCTGGACCCC
     1201 CCCGGCGGAG GAGACGGCCT TGGTTTGGGA CACGAGAAGT TCCCCAGCGA CCTGGACCTG
     1261 GACATGTTCA ACGGCAGCCT GGAGTGCGAC ATGGAGAGCA TCATCAGAAG CGAGCTGATG
     1321 GACGCCGACG GCCTGGACTT CAACTTCGAC AGCtaggccg ctcgaggagg gcccgaacaa
     1381 aaactcatct cagaagagga tctgaatagc gccgtcgacc atcatcataa tcatcattga
     1441 gtttaaaccc gctgatcagc ctcgactgtg ccttctagtt gccagccatc tgttgtttgc
     1501 ccctcccccg tgccttcctt gaccctggaa ggtgccactc ccactgtcct ttcctaataa
     1561 aatgaggaaa ttgcatcgca ttgtctgagt aggtgtcatt ctattctggg gggtggggtg
     1621 gggcaggaca gcaaggggga ggattgggaa gacaatagca ggcatgctgg ggatgcggtg
     1681 ggctctatgg agatctatcg atgttctggg gatgcggtgg gctctatggc ttctgaggcg
     1741 gaaagaacca gctggggctc tagggggtat ccccacgcgc cctgtagcgg cgcattaagc
     1801 gcggcgggtg tggtggttac gcgcagcgtg accgctacac ttgccagcgc cctagcgccc
     1861 gctcctttcg ctttcttccc ttcctttctc gccacgttcg ccggctttcc ccgtcaagct
     1921 ctaaatcggg ggctcccttt agggttccga tttagtgctt tacggcacct cgaccccaaa
     1981 aaacttgatt agggtgatgg ttcacgtacc tagaagttcc tattccgaag ttcctattct
     2041 ctagaaagta taggaacttc cttggccaaa aagcctgaac tcaccgcgac gtctgtcgag
     2101 aagtttctga tcgaaaagtt cgacagcgtc tccgacctga tgcagctctc ggagggcgaa
     2161 gaatctcgtg ctttcagctt cgatgtagga gggcgtggat atgtcctgcg ggtaaatagc
     2221 tgcgccgatg gtttctacaa agatcgttat gtttatcggc actttgcatc ggccgcgctc
     2281 ccgattccgg aagtgcttga cattggggaa ttcagcgaga gcctgaccta ttgcatctcc
     2341 cgccgtgcac agggtgtcac gttgcaagac ctgcctgaaa ccgaactgcc cgctgttctg
     2401 cagccggtcg cggaggccat ggatgcgatc gctgcggccg atcttagcca gacgagcggg
     2461 ttcggcccat tcggaccgca aggaatcggt caatacacta catggcgtga tttcatatgc
     2521 gcgattgctg atccccatgt gtatcactgg caaactgtga tggacgacac cgtcagtgcg
     2581 tccgtcgcgc aggctctcga tgagctgatg ctttgggccg aggactgccc cgaagtccgg
     2641 cacctcgtgc acgcggattt cggctccaac aatgtcctga cggacaatgg ccgcataaca
     2701 gcggtcattg actggagcga ggcgatgttc ggggattccc aatacgaggt cgccaacatc
     2761 ttcttctgga ggccgtggtt ggcttgtatg gagcagcaga cgcgctactt cgagcggagg
     2821 catccggagc ttgcaggatc gccgcggctc cgggcgtata tgctccgcat tggtcttgac
     2881 caactctatc agagcttggt tgacggcaat ttcgatgatg cagcttgggc gcagggtcga
     2941 tgcgacgcaa tcgtccgatc cggagccggg actgtcgggc gtacacaaat cgcccgcaga
     3001 agcgcggccg tctggaccga tggctgtgta gaagtactcg ccgatagtgg aaaccgacgc
     3061 cccagcactc gtccgagggc aaaggaatag cacgtactac gagatttcga ttccaccgcc
     3121 gccttctatg aaaggttggg cttcggaatc gttttccggg acgccggctg gatgatcctc
     3181 cagcgcgggg atctcatgct ggagttcttc gcccacccca acttgtttat tgcagcttat
     3241 aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg
     3301 cattctagtt gtggtttgtc caaactcatc aatgtatctt atcatgtctg tataccgtcg
     3361 acctctagct agagcttggc gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat
     3421 ccgctcacaa ttccacacaa catacgagcc ggaagcataa agtgtaaagc ctggggtgcc
     3481 taatgagtga gctaactcac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga
     3541 aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt
     3601 attgggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg
     3661 cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac
     3721 gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg
     3781 ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca
     3841 agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc
     3901 tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc
     3961 ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag
     4021 gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc
     4081 ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca
     4141 gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg
     4201 aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg
     4261 aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct
     4321 ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa
     4381 gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa
     4441 gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa
     4501 tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc
     4561 ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga
     4621 ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca
     4681 atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc
     4741 ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat
     4801 tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc
     4861 attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt
     4921 tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc
     4981 ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg
     5041 gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt
     5101 gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg
     5161 gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga
     5221 aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg
     5281 taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg
     5341 tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt
     5401 tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc
     5461 atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca
     5521 tttccccgaa aagtgccacc tgacgtcgac ggatcgggag atctcccgat cccctatggt
     5581 gcactctcag tacaatctgc tctgatgccg catagttaag ccagtatctg ctccctgctt
     5641 gtgtgttgga ggtcgctgag tagtgcgcga gcaaaattta agctacaaca aggcaaggct
     5701 tgaccgacaa ttgcatgaag aatctgctta gggttaggcg ttttgcgctg cttcgtacgg
     5761 gccagata
//