LOCUS       AAV-U6-sgPlxnb2-KO-2-EF 5592 bp ds-DNA     circular     15-JUN-2022
DEFINITION  .
FEATURES             Location/Qualifiers
     repeat_region   complement(1..141)
                     /label="AAV2 ITR"
                     /ApEinfo_revcolor="#ff9ccd"
                     /ApEinfo_fwdcolor="#ff9ccd"
                     /note="note: pLannotate database: snapgene identity: 100.0 match_length: 100.0 fragment: False other: repeat_region"
     primer          140..177
                     /label="InF-U6-F"
                     /note="sequence: ctgcggcccccgttagagggcctatttcccatgattcc"
                     /ApEinfo_revcolor="#85dae9"
                     /ApEinfo_fwdcolor="#85dae9"
     promoter        155..395
                     /label="U6 promoter"
                     /ApEinfo_revcolor="#faac61"
                     /ApEinfo_fwdcolor="#faac61"
                     /note="note: pLannotate database: snapgene identity: 100.0 match_length: 100.0 fragment: False other: promoter"
     misc_feature    201..1318
                     /label="sanger (U6-f)"
                     /ApEinfo_revcolor="#75c6a9"
                     /ApEinfo_fwdcolor="#75c6a9"
     primer          400..423
                     /label="AAV-sgPlxnb2-2-EF1a-LibVec FWD"
                     /note="sequence: caccGCAGTACTACGAGTGTCGGG"
                     /ApEinfo_revcolor="#f58a5e"
                     /ApEinfo_fwdcolor="#f58a5e"
     misc_feature    403..403
                     /label="del. (it should not affect the expression of the sgRNA)"
                     /ApEinfo_revcolor="#b4abac"
                     /ApEinfo_fwdcolor="#b4abac"
     misc_feature    404..423
                     /label="Guide"
                     /ApEinfo_revcolor="#7ea7d8"
                     /ApEinfo_fwdcolor="#7ea7d8"
     primer          complement(404..427)
                     /label="AAV-sgPlxnb2-2-EF1a-LibVec REV"
                     /note="sequence: aaacCCCGACACTCGTAGTACTGC"
                     /ApEinfo_revcolor="#75c6a9"
                     /ApEinfo_fwdcolor="#75c6a9"
     misc_RNA        424..499
                     /label="gRNA scaffold"
                     /ApEinfo_revcolor="#85dae9"
                     /ApEinfo_fwdcolor="#85dae9"
                     /note="note: pLannotate database: snapgene identity: 100.0 match_length: 100.0 fragment: False other: misc_RNA"
     primer          474..492
                     /label="qPCR-gRNA-F"
                     /note="sequence: ttgaaaaagtggcaccgag"
                     /ApEinfo_revcolor="#b7e6d7"
                     /ApEinfo_fwdcolor="#b7e6d7"
     primer          complement(485..520)
                     /label="InF-gRNA-R"
                     /note="sequence: tccatctttgcattaaaaaaagcaccgactcggtgc"
                     /ApEinfo_revcolor="#b1ff67"
                     /ApEinfo_fwdcolor="#b1ff67"
     CDS             516..582
                     /label="small t antigen (fragment)"
                     /ApEinfo_revcolor="#b1ff67"
                     /ApEinfo_fwdcolor="#b1ff67"
                     /note="note: pLannotate database: snapgene identity: 100.0 match_length: 12.8 fragment: True other: CDS"
     CDS             516..582
                     /ApEinfo_revcolor="#84b0dc"
                     /ApEinfo_fwdcolor="#84b0dc"
                     /translation="MDKVLNREESLQLMDLLGLERS"
     primer          complement(567..586)
                     /label="qPCR-EF1a-R"
                     /note="sequence: ttcccactcctttcaagacc"
                     /ApEinfo_revcolor="#75c6a9"
                     /ApEinfo_fwdcolor="#75c6a9"
     promoter        589..1767
                     /label="EF-1α promoter"
                     /ApEinfo_revcolor="#f8d3a9"
                     /ApEinfo_fwdcolor="#f8d3a9"
                     /note="note: pLannotate database: snapgene identity: 100.0 match_length: 100.0 fragment: False other: promoter"
     CDS             1781..2500
                     /label="EGFP"
                     /ApEinfo_revcolor="#f8d3a9"
                     /ApEinfo_fwdcolor="#f8d3a9"
                     /note="note: pLannotate database: snapgene identity: 100.0 match_length: 100.0 fragment: False other: CDS"
     CDS             1781..2500
                     /ApEinfo_revcolor="#84b0dc"
                     /ApEinfo_fwdcolor="#84b0dc"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDGNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKVNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLEFVTAAGITLGMDELYK*"
     promoter        complement(2568..2586)
                     /label="SP6 promoter"
                     /ApEinfo_revcolor="#b1ff67"
                     /ApEinfo_fwdcolor="#b1ff67"
                     /note="note: pLannotate database: snapgene identity: 100.0 match_length: 100.0 fragment: False other: promoter"
     polyA_signal    2612..2836
                     /label="bGH poly(A) signal"
                     /ApEinfo_revcolor="#b4abac"
                     /ApEinfo_fwdcolor="#b4abac"
                     /note="note: pLannotate database: snapgene identity: 100.0 match_length: 100.0 fragment: False other: polyA_signal"
     repeat_region   2855..2995
                     /label="AAV2 ITR"
                     /ApEinfo_revcolor="#b7e6d7"
                     /ApEinfo_fwdcolor="#b7e6d7"
                     /note="note: pLannotate database: snapgene identity: 100.0 match_length: 100.0 fragment: False other: repeat_region"
     misc_feature    complement(3000..3051)
                     /label="bom (fragment)"
                     /ApEinfo_revcolor="#b1ff67"
                     /ApEinfo_fwdcolor="#b1ff67"
                     /note="note: pLannotate database: snapgene identity: 100.0 match_length: 36.9 fragment: True other: misc_feature"
     rep_origin      3070..3450
                     /label="M13 ori"
                     /ApEinfo_revcolor="#ffef86"
                     /ApEinfo_fwdcolor="#ffef86"
                     /note="note: pLannotate database: snapgene identity: 100.0 match_length: 100.0 fragment: False other: rep_origin"
     misc_feature    complement(3526..3575)
                     /label="bom (fragment)"
                     /ApEinfo_revcolor="#c7b0e3"
                     /ApEinfo_fwdcolor="#c7b0e3"
                     /note="note: pLannotate database: snapgene identity: 100.0 match_length: 35.5 fragment: True other: misc_feature"
     CDS             complement(3677..3711)
                     /label="rop (fragment)"
                     /ApEinfo_revcolor="#c6c9d1"
                     /ApEinfo_fwdcolor="#c6c9d1"
                     /note="note: pLannotate database: snapgene identity: 100.0 match_length: 18.2 fragment: True other: CDS"
     CDS             complement(3677..3711)
                     /ApEinfo_revcolor="#84b0dc"
                     /ApEinfo_fwdcolor="#84b0dc"
                     /translation="SRVSVMTVKTS"
     promoter        3807..3911
                     /label="AmpR promoter"
                     /ApEinfo_revcolor="#ffef86"
                     /ApEinfo_fwdcolor="#ffef86"
                     /note="note: pLannotate database: snapgene identity: 100.0 match_length: 100.0 fragment: False other: promoter"
     CDS             3912..4772
                     /label="AmpR"
                     /ApEinfo_revcolor="#c7b0e3"
                     /ApEinfo_fwdcolor="#c7b0e3"
                     /note="note: pLannotate database: snapgene identity: 99.8 match_length: 100.0 fragment: False other: CDS"
     CDS             3912..4772
                     /ApEinfo_revcolor="#84b0dc"
                     /ApEinfo_fwdcolor="#84b0dc"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYIELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYSPVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRWEPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSALPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGASLIKHW*"
     rep_origin      4943..5531
                     /label="ori"
                     /ApEinfo_revcolor="#ff9ccd"
                     /ApEinfo_fwdcolor="#ff9ccd"
                     /note="note: pLannotate database: snapgene identity: 100.0 match_length: 100.0 fragment: False other: rep_origin"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcaaag cccgggcgtc
       61 gggcgacctt tggtcgcccg gcctcagtga gcgagcgagc gcgcagagag ggagtggcca
      121 actccatcac taggggttcc tgcggccccc gttagagggc ctatttccca tgattccttc
      181 atatttgcat atacgataca aggctgttag agagataatt agaattaatt tgactgtaaa
      241 cacaaagata ttagtacaaa atacgtgacg tagaaagtaa taatttcttg ggtagtttgc
      301 agttttaaaa ttatgtttta aaatggacta tcatatgctt accgtaactt gaaagtattt
      361 cgatttcttg gctttatata tcttgtggaa aggacgaaac accGCAGTAC TACGAGTGTC
      421 GGGgttttag agctagaaat agcaagttaa aataaggcta gtccgttatc aacttgaaaa
      481 agtggcaccg agtcggtgct ttttttaatg caaagatgga taaagtttta aacagagagg
      541 aatctttgca gctaatggac cttctaggtc ttgaaaggag tgggaattgg ctccggtgcc
      601 cgtcagtggg cagagcgcac atcgcccaca gtccccgaga agttgggggg aggggtcggc
      661 aattgaaccg gtgcctagag aaggtggcgc ggggtaaact gggaaagtga tgtcgtgtac
      721 tggctccgcc tttttcccga gggtggggga gaaccgtata taagtgcagt agtcgccgtg
      781 aacgttcttt ttcgcaacgg gtttgccgcc agaacacagg taagtgccgt gtgtggttcc
      841 cgcgggcctg gcctctttac gggttatggc ccttgcgtgc cttgaattac ttccacctgg
      901 ctgcagtacg tgattcttga tcccgagctt cgggttggaa gtgggtggga gagttcgagg
      961 ccttgcgctt aaggagcccc ttcgcctcgt gcttgagttg aggcctggcc tgggcgctgg
     1021 ggccgccgcg tgcgaatctg gtggcacctt cgcgcctgtc tcgctgcttt cgataagtct
     1081 ctagccattt aaaatttttg atgacctgct gcgacgcttt ttttctggca agatagtctt
     1141 gtaaatgcgg gccaagatct gcacactggt atttcggttt ttggggccgc gggcggcgac
     1201 ggggcccgtg cgtcccagcg cacatgttcg gcgaggcggg gcctgcgagc gcggccaccg
     1261 agaatcggac gggggtagtc tcaagctggc cggcctgctc tggtgcctgg cctcgcgccg
     1321 ccgtgtatcg ccccgccctg ggcggcaagg ctggcccggt cggcaccagt tgcgtgagcg
     1381 gaaagatggc cgcttcccgg ccctgctgca gggagctcaa aatggaggac gcggcgctcg
     1441 ggagagcggg cgggtgagtc acccacacaa aggaaaaggg cctttccgtc ctcagccgtc
     1501 gcttcatgtg actccacgga gtaccgggcg ccgtccaggc acctcgatta gttctcgagc
     1561 ttttggagta cgtcgtcttt aggttggggg gaggggtttt atgcgatgga gtttccccac
     1621 actgagtggg tggagactga agttaggcca gcttggcact tgatgtaatt ctccttggaa
     1681 tttgcccttt ttgagtttgg atcttggttc attctcaagc ctcagacagt ggttcaaagt
     1741 ttttttcttc catttcaggt gtcgtgagcg gccgctcgag atggtgagca agggcgagga
     1801 gctgttcacc ggggtggtgc ccatcctggt cgagctggac ggcgacgtaa acggccacaa
     1861 gttcagcgtg tccggcgagg gcgagggcga tgccacctac ggcaagctga ccctgaagtt
     1921 catctgcacc accggcaagc tgcccgtgcc ctggcccacc ctcgtgacca ccctgaccta
     1981 cggcgtgcag tgcttcagcc gctaccccga ccacatgaag cagcacgact tcttcaagtc
     2041 cgccatgccc gaaggctacg tccaggagcg caccatcttc ttcaaggacg acggcaacta
     2101 caagacccgc gccgaggtga agttcgaggg cgacaccctg gtgaaccgca tcgagctgaa
     2161 gggcatcgac ttcaaggagg acggcaacat cctggggcac aagctggagt acaactacaa
     2221 cagccacaac gtctatatca tggccgacaa gcagaagaac ggcatcaagg tgaacttcaa
     2281 gatccgccac aacatcgagg acggcagcgt gcagctcgcc gaccactacc agcagaacac
     2341 ccccatcggc gacggccccg tgctgctgcc cgacaaccac tacctgagca cccagtccgc
     2401 cctgagcaaa gaccccaacg agaagcgcga tcacatggtc ctgctggagt tcgtgaccgc
     2461 cgccgggatc actctcggca tggacgagct gtacaagtaa gagcgcaccc gtccgagcta
     2521 aggcgcgcca tacatgcatg gtgtacaagt aatctagagg gccctattct atagtgtcac
     2581 ctaaatgcta gagctcgctg atcagcctcg actgtgcctt ctagttgcca gccatctgtt
     2641 gtttgcccct cccccgtgcc ttccttgacc ctggaaggtg ccactcccac tgtcctttcc
     2701 taataaaatg aggaaattgc atcgcattgt ctgagtaggt gtcattctat tctggggggt
     2761 ggggtggggc aggacagcaa gggggaggat tgggaagaca atagcaggca tgctggggat
     2821 gcggtgggct ctatggcttc tgaggcgggg ccgcaggaac ccctagtgat ggagttggcc
     2881 actccctctc tgcgcgctcg ctcgctcact gaggccgggc gaccaaaggt cgcccgacgc
     2941 ccgggctttg cccgggcggc ctcagtgagc gagcgagcgc gcagctgcct gcaggggcgc
     3001 ctgatgcggt attttctcct tacgcatctg tgcggtattt cacaccgcat acgtcaaagc
     3061 aaccatagta cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca
     3121 gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc tttcgctttc ttcccttcct
     3181 ttctcgccac gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt
     3241 tccgatttag tgctttacgg cacctcgacc ccaaaaaact tgatttgggt gatggttcac
     3301 gtagtgggcc atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct
     3361 ttaatagtgg actcttgttc caaactggaa caacactcaa ccctatctcg ggctattctt
     3421 ttgatttata agggattttg ccgatttcgg cctattggtt aaaaaatgag ctgatttaac
     3481 aaaaatttaa cgcgaatttt aacaaaatat taacgtttac aattttatgg tgcactctca
     3541 gtacaatctg ctctgatgcc gcatagttaa gccagccccg acacccgcca acacccgctg
     3601 acgcgccctg acgggcttgt ctgctcccgg catccgctta cagacaagct gtgaccttct
     3661 ccgggagctg catgtgtcag aggttttcac cgtcatcacc gaaacgcgcg atacgaaagg
     3721 gcctcgtgat acgcctattt ttataggtta atgtcatgat aataatggtt tcttagacgt
     3781 caggtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac
     3841 attcaaatat gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa
     3901 aaaggaagag tatgagtatt caacatttcc gtgtcgccct tattcccttt tttgcggcat
     3961 tttgccttcc tgtttttgct cacccagaaa cgctggtgaa agtaaaagat gctgaagatc
     4021 agttgggtgc acgagtgggt tacatcgaac tggatctcaa cagcggtaag atccttgaga
     4081 gttttcgccc cgaagaacgt tttccaatga tgagcacttt taaagttctg ctatgtggcg
     4141 cggtattatc ccgtattgac gccgggcaag agcaactcgg tcgccgcata cactattctc
     4201 agaatgactt ggttgagtac tcaccagtca cagaaaagca tcttacggat ggcatgacag
     4261 taagagaatt atgcagtgct gccataacca tgagtgataa cactgcggcc aacttacttc
     4321 tgacaacgat cggaggaccg aaggagctaa ccgctttttt gcacaacatg ggggatcatg
     4381 taactcgcct tgatcgttgg gaaccggagc tgaatgaagc cataccaaac gacgagcgtg
     4441 acaccacgat gcctgtagca atggcaacaa cgttgcgcaa actattaact ggcgaactac
     4501 ttactctagc ttcccggcaa caattaatag actggatgga ggcggataaa gttgcaggac
     4561 cacttctgcg ctcggccctt ccggctggct ggtttattgc tgataaatct ggagccggtg
     4621 agcgtgggtc tcgcggtatc attgcagcac tggggccaga tggtaagccc tcccgtatcg
     4681 tagttatcta cacgacgggg agtcaggcaa ctatggatga acgaaataga cagatcgctg
     4741 agataggtgc ctcactgatt aagcattggt aactgtcaga ccaagtttac tcatatatac
     4801 tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg
     4861 ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg
     4921 tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc
     4981 aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc
     5041 tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtc cttctagtgt
     5101 agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc
     5161 taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact
     5221 caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac
     5281 agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag
     5341 aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg
     5401 gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg
     5461 tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga
     5521 gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt
     5581 ttgctcacat gt
//