LOCUS       pCL026_AAVS1-puroR-9xTe 7914 bp ds-DNA     circular     16-NOV-2021
DEFINITION  .
COMMENT     reporter for high-throughput recruitment libraries integrate into
            AAVs1 locus Expression screening: use Citrine and myc-mag
            separation HDR screening: use Cas9 knock-in of FLAG, or knockout
            citrine/myc for NHEJ
FEATURES             Location/Qualifiers
     protein_bind    819..840
                     /label="CAP binding site"
     promoter        855..860
                     /label="lac promoter"
     promoter        861..878
                     /label="lac promoter"
     promoter        879..885
                     /label="lac promoter"
     protein_bind    893..909
                     /label="lac operator"
     primer_bind     917..933
                     /label="M13 rev"
     promoter        954..972
                     /label="T3 promoter"
     misc_feature    1007..1810
                     /label="left_arm"
     misc_feature    1817..1842
                     /label="SA"
     CDS             1866..1919
                     /label="T2A"
     CDS             1866..1928
                     /label="Translation 1866-1928"
                     /translation="EGRGSLLTCGDVEENPGPRLE"
     CDS             1929..2528
                     /label="PuroR"
     CDS             1929..2528
                     /label="Translation 1929-2528"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIERVTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLAAQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETSAPRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA*"
     misc_feature    2566..2790
                     /label="polyA"
     primer          complement(2635..2654)
                     /label="QB3668"
                     /note="sequence: ggaaaggacagtgggagtgg"
     misc_feature    2845..3168
                     /label="9xTRE"
     misc_feature    2862..2880
                     /label="TRE"
     misc_feature    2898..2916
                     /label="TRE"
     misc_feature    2934..2952
                     /label="TRE"
     misc_feature    2970..2988
                     /label="TRE"
     misc_feature    3006..3024
                     /label="TRE"
     misc_feature    3042..3060
                     /label="TRE"
     misc_feature    3078..3096
                     /label="TRE"
     misc_feature    3114..3132
                     /label="TRE"
     misc_feature    3150..3168
                     /label="TRE"
     primer          3178..3223
                     /label="oCL021"
                     /note="sequence: caggtagggctgtacagtgggaatgcatggggttggggttgcgcct"
     misc_feature    3206..3716
                     /label="phPGK"
     misc_feature    3275..3284
                     /label="STAT4 mouse"
     misc_feature    3456..3461
                     /label="old BsmBI site"
     primer          complement(3694..3744)
                     /label="oCL022"
                     /note="sequence: CTGTCTCCATggtggcccaccgcggtggctggggagagaggtcggtgattc"
     regulatory      3729..3734
                     /label="Kozak sequence"
     CDS             3735..3797
                     /label="Igκ leader"
     misc_feature    3785..3804
                     /label="sgRNA_Sp"
     misc_feature    3791..3810
                     /label="sgRNA_Sa"
     CDS             3837..4517
                     /label="hIgG1-Fc"
     CDS             4518..4565
                     /label="Linker"
     CDS             4566..4595
                     /label="Myc"
     CDS             4596..4745
                     /label="PDGFRB Transmembrane Domain"
     CDS             4761..4814
                     /label="T2A"
     regulatory      4821..4826
                     /label="Kozak sequence"
     CDS             4827..4829
                     /label="Citrine"
     CDS             4827..5564
                     /label="Translation 4827-5564"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKFICTTGKLPVPWPTLVTTFGYGLMCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDGNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKVNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLLEFVTAAGITLGMDELYKPKKKRKV"
     CDS             4830..4832
                     /label="Citrine"
     CDS             4833..5543
                     /label="Citrine"
     misc_feature    4872..4893
                     /label="QG0021"
     CDS             5544..5564
                     /label="SV40 NLS"
     CDS             5565..5567
                     /label="Translation 5565-5567"
                     /translation="*"
     misc_feature    5570..5573
                     /label="SV40 pA"
     misc_feature    5574..5577
                     /label="SV40 pA"
     polyA_signal    5575..5696
                     /label="SV40 poly(A) signal"
     misc_feature    5578..5578
                     /label="SV40 pA"
     misc_feature    5579..5579
                     /label="SV40 pA"
     misc_feature    5580..5705
                     /label="SV40 pA"
     misc_feature    5739..5746
                     /label="Insertion from Gibson assembly"
     misc_feature    5762..6598
                     /label="right_arm"
     CDS             complement(6873..7664)
                     /label="AmpR"
     CDS             complement(7665..7733)
                     /label="AmpR"
     rep_origin      7857..531
                     /label="ori"
ORIGIN
        1 ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc
       61 ttcagcagag cgcagatacc aaatactgtt cttctagtgt agccgtagtt aggccaccac
      121 ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct
      181 gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat
      241 aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg
      301 acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa
      361 gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg
      421 gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga
      481 cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc
      541 aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat gttctttcct
      601 gcgttatccc ctgattctgt ggataaccgt attaccgcct ttgagtgagc tgataccgct
      661 cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg aggaagcgga agagcgccca
      721 atacgcaaac cgcctctccc cgcgcgttgg ccgattcatt aatgcagctg gcacgacagg
      781 tttcccgact ggaaagcggg cagtgagcgc aacgcaatta atgtgagtta gctcactcat
      841 taggcacccc aggctttaca ctttatgctt ccggctcgta tgttgtgtgg aattgtgagc
      901 ggataacaat ttcacacagg aaacagctat gaccatgatt acgccaagct cagaattaac
      961 cctcactaaa gggactagtc ctgcaggttt aaacgaattc gccctttgct ttctctgacc
     1021 agcattctct cccctgggcc tgtgccgctt tctgtctgca gcttgtggcc tgggtcacct
     1081 ctacggctgg cccagatcct tccctgccgc ctccttcagg ttccgtcttc ctccactccc
     1141 tcttcccctt gctctctgct gtgttgctgc ccaaggatgc tctttccgga gcacttcctt
     1201 ctcggcgctg caccacgtga tgtcctctga gcggatcctc cccgtgtctg ggtcctctcc
     1261 gggcatctct cctccctcac ccaaccccat gccgtcttca ctcgctgggt tcccttttcc
     1321 ttctccttct ggggcctgtg ccatctctcg tttcttagga tggccttctc cgacggatgt
     1381 ctcccttgcg tcccgcctcc ccttcttgta ggcctgcatc atcaccgttt ttctggacaa
     1441 ccccaaagta ccccgtctcc ctggctttag ccacctctcc atcctcttgc tttctttgcc
     1501 tggacacccc gttctcctgt ggattcgggt cacctctcac tcctttcatt tgggcagctc
     1561 ccctaccccc cttacctctc tagtctgtgc tagctcttcc agccccctgt catggcatct
     1621 tccaggggtc cgagagctca gctagtcttc ttcctccaac ccgggcccct atgtccactt
     1681 caggacagca tgtttgctgc ctccagggat cctgtgtccc cgagctggga ccaccttata
     1741 ttcccagggc cggttaatgt ggctctggtt ctgggtactt ttatctgtcc cctccacccc
     1801 acagtggggc aagcttctga cctcttctct tcctcccaca gggcctcgag agatctggca
     1861 gcggagaggg cagaggaagt cttctaacat gcggtgacgt ggaggagaat cccggcccta
     1921 ggctcgagat gaccgagtac aagcccacgg tgcgcctcgc cacccgcgac gacgtcccca
     1981 gggccgtacg caccctcgcc gccgcgttcg ccgactaccc cgccacgcgc cacaccgtcg
     2041 atccggaccg ccacatcgag cgggtcaccg agctgcaaga actcttcctc acgcgcgtcg
     2101 ggctcgacat cggcaaggtg tgggtcgcgg acgacggcgc cgcggtggcg gtctggacca
     2161 cgccggagag cgtcgaagcg ggggcggtgt tcgccgagat cggcccgcgc atggccgagt
     2221 tgagcggttc ccggctggcc gcgcagcaac agatggaagg cctcctggcg ccgcaccggc
     2281 ccaaggagcc cgcgtggttc ctggccaccg tcggcgtctc gcccgaccac cagggcaagg
     2341 gtctgggcag cgccgtcgtg ctccccggag tggaggcggc cgagcgcgcc ggggtgcccg
     2401 ccttcctgga gacctccgcg ccccgcaacc tccccttcta cgagcggctc ggcttcaccg
     2461 tcaccgccga cgtcgaggtg cccgaaggac cgcgcacctg gtgcatgacc cgcaagcccg
     2521 gtgcctgatc tagagggccc gtttaaaccc gctgatcagc ctcgactgtg ccttctagtt
     2581 gccagccatc tgttgtttgc ccctcccccg tgccttcctt gaccctggaa ggtgccactc
     2641 ccactgtcct ttcctaataa aatgaggaaa ttgcatcgca ttgtctgagt aggtgtcatt
     2701 ctattctggg gggtggggtg gggcaggaca gcaaggggga ggattgggaa gacaatagca
     2761 ggcatgctgg ggatgcggtg ggctctatgg cttctgaggc ggaagaacca gctgggggct
     2821 agcaaagcct aggaacagct gcatatagca ctgcagttta ctccctatca gtgatagaga
     2881 gtcaaaaggc catgccctcc ctatcagtga tagagagagc agactcaagg tggtccctat
     2941 cagtgataga gagattgagc aagagggcct ccctatcagt gatagagaat agcactgcag
     3001 tttactccct atcagtgata gagagtcaaa aggccatgcc ctccctatca gtgatagaga
     3061 gagcagactc aaggtggtcc ctatcagtga tagagagatt gagcaagagg gcctccctat
     3121 cagtgataga gattgatcca gtcagggcct ccctatcagt gatagagaag ctatgtgcag
     3181 gtagggctgt acagtgggaa tgcatggggt tggggttgcg ccttttccaa ggcagccctg
     3241 ggtttgcgca gggacgcggc tgctctgggc gtggttccgg gaaacgcagc ggcgccgacc
     3301 ctgggtctcg cacattcttc acgtccgttc gcagcgtcac ccggatcttc gccgctaccc
     3361 ttgtgggccc cccggcgacg cttcctgctc cgcccctaag tcgggaaggt tccttgcggt
     3421 tcgcggcgtg ccggacgtga caaacggaag ccgcacgtAt cactagtacc ctcgcagacg
     3481 gacagcgcca gggagcaatg gcagcgcgcc gaccgcgatg ggctgtggcc aatagcggct
     3541 gctcagcagg gcgcgccgag agcagcggcc gggaaggggc ggtgcgggag gcggggtgtg
     3601 gggcggtagt gtgggccctg ttcctgcccg cgcggtgttc cgcattctgc aagcctccgg
     3661 agcgcacgtc ggcagtcggc tccctcgttg accgaatcac cgacctctct ccccagccac
     3721 cgcggtgggc caccATGGAG ACAGACACAC TCCTGCTATG GGTACTGCTG CTCTGGGTTC
     3781 CAGGTTCCAC TGGTGACGCT AGCGAGACGG GCGGATCAGG CGGTCGTCTC GGATCCGACA
     3841 AAACTCACAC ATGCCCACCG TGCCCAGCAC CTGAACTCCT GGGGGGACCG TCAGTCTTCC
     3901 TCTTCCCCCC AAAACCCAAG GACACCCTCA TGATCTCCCG GACCCCTGAG GTCACATGCG
     3961 TGGTGGTGGA CGTGAGCCAC GAAGACCCTG AGGTCAAGTT CAACTGGTAC GTGGACGGCG
     4021 TGGAGGTGCA TAATGCCAAG ACAAAGCCGC GGGAGGAGCA GTACAACAGC ACGTACCGTG
     4081 TGGTCAGCGT CCTCACCGTC CTGCACCAGG ACTGGCTGAA TGGCAAGGAG TACAAGTGCA
     4141 AGGTCTCCAA CAAAGCCCTC CCAGCCCCCA TCGAGAAAAC CATCTCCAAA GCCAAAGGGC
     4201 AGCCCCGAGA ACCACAGGTC TACACCCTGC CCCCATCCCG GGAGGAGATG ACCAAGAACC
     4261 AGGTCAGCCT GACCTGCCTG GTCAAAGGCT TCTATCCCAG CGACATCGCC GTGGAGTGGG
     4321 AGAGCAATGG GCAGCCGGAG AACAACTACA AGACCACGCC TCCCGTGCTG GACTCCGACG
     4381 GCTCCTTCTT CCTCTACAGC AAGCTCACCG TGGACAAGAG CAGGTGGCAG CAGGGGAACG
     4441 TCTTCTCATG CTCCGTGATG CACGAGGCTC TGCACAACCA CTACACGCAG AAGAGCCTCT
     4501 CCCTGTCTCC GGGTAAAGGA GGAGGTGGAA GCGGAGGAGG AGGAAGCGGA GGAGGAGGTA
     4561 GCggaGAACA AAAACTCATC TCAGAAGAGG ATCTGAATGC TGTGGGCCAG GACACGCAGG
     4621 AGGTCATCGT GGTGCCACAC TCCTTGCCCT TTAAGGTGGT GGTGATCTCA GCCATCCTGG
     4681 CCCTGGTGGT GCTCACCATC ATCTCCCTTA TCATCCTCAT CATGCTTTGG CAGAAGAAGC
     4741 CACGTtgtac aGGCAGTGGA GAGGGCAGAG GAAGTCTGCT AACATGCGGT GACGTCGAGG
     4801 AGAATCCTGG CCCAccggtc gccaccatgg tgagcaaggg cgaggagctg ttcaccgggg
     4861 tggtgcccat cctggtcgag ctggacggcg acgtaaacgg ccacaagttc agcgtgtccg
     4921 gcgagggcga gggcgatgcc acctacggca agctgaccct gaagttcatc tgcaccaccg
     4981 gcaagctgcc cgtgccctgg cccaccctcg tgaccacctt cggctacggc ctgatgtgct
     5041 tcgcccgcta ccccgaccac atgaagcagc acgacttctt caagtccgcc atgcccgaag
     5101 gctacgtcca ggagcgcacc atcttcttca aggacgacgg caactacaag acccgcgccg
     5161 aggtgaagtt cgagggcgac accctggtga accgcatcga gctgaagggc atcgacttca
     5221 aggaggacgg caacatcctg gggcacaagc tggagtacaa ctacaacagc cacaacgtct
     5281 atatcatggc cgacaagcag aagaacggca tcaaggtgaa cttcaagatc cgccacaaca
     5341 tcgaggacgg cagcgtgcag ctcgccgacc actaccagca gaacaccccc atcggcgacg
     5401 gccccgtgct gctgcccgac aaccactacc tgagctacca gtccgccctg agcaaagacc
     5461 ccaacgagaa gcgcgatcac atggtcctgc tggagttcgt gaccgccgcc gggatcactc
     5521 tcggcatgga cgagctgtac aagCCAAAAA AGAAGAGAAA GGTAtagtta actaaacttg
     5581 tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa
     5641 gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttatcat
     5701 gtctgCAtat gtaaattgta agcgtctcgg cctctagaGT CGAGATgaca ttgattattg
     5761 aactagggac aggattggtg acagaaaagc cccatcctta ggcctcctcc ttcctagtct
     5821 cctgatattg ggtctaaccc ccacctcctg ttaggcagat tccttatctg gtgacacacc
     5881 cccatttcct ggagccatct ctctccttgc cagaacctct aaggtttgct tacgatggag
     5941 ccagagagga tcctgggagg gagagcttgg cagggggtgg gagggaaggg ggggatgcgt
     6001 gacctgcccg gttctcagtg gccaccctgc gctaccctct cccagaacct gagctgctct
     6061 gacgcggctg tctggtgcgt ttcactgatc ctggtgctgc agcttcctta cacttcccaa
     6121 gaggagaagc agtttggaaa aacaaaatca gaataagttg gtcctgagtt ctaactttgg
     6181 ctcttcacct ttctagtccc caatttatat tgttcctccg tgcgtcagtt ttacctgtga
     6241 gataaggcca gtagccagcc ccgtcctggc agggctgtgg tgaggagggg ggtgtccgtg
     6301 tggaaaactc cctttgtgag aatggtgcgt cctaggtgtt caccaggtcg tggccgcctc
     6361 tactcccttt ctctttctcc atccttcttt ccttaaagag tccccagtgc tatctgggac
     6421 atattcctcc gcccagagca gggtcccgct tccctaaggc cctgctctgg gcttctgggt
     6481 ttgagtcctt ggcaagccca ggagaggcgc tcaggcttcc ctgtccccct tcctcgtcca
     6541 ccatctcatg cccctggctc tcctgcccct tccctacagg ggttcctggc tctgctcttc
     6601 gccttcttga cgagttcttc tgaattatta acgcttacaa tttcctgatg cggtattttc
     6661 tccttacgca tctgtgcggt atttcacacc gcatcaggtg gcacttttcg gggaaatgtg
     6721 cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga
     6781 ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc
     6841 taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct
     6901 atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata
     6961 actacgatac gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca
     7021 cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga
     7081 agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga
     7141 gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg
     7201 gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga
     7261 gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt
     7321 gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct
     7381 cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca
     7441 ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat
     7501 accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga
     7561 aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc
     7621 aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg
     7681 caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc
     7741 ctttttcaat attattgaag catttatcag ggttattgtc tcatgaccaa aatcccttaa
     7801 cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga
     7861 gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gcta
//