LOCUS       AAV-U6-sgRNA-EF1a-LibVe 7453 bp ds-DNA     circular     23-MAY-2022
DEFINITION  .
COMMENT     Imported using the Genbank importer. File name: 315630_pLann.gbk
            Annotated with pLannotate v1.2.0 Linear. AAV-EF1a-LibVec with PacI
            -> InFusion with amplicon Template: pT4-U6-sgRNA-CMV-EGFP InF-U6-F:
            ctgcggcccccgttagagggcctatttcccatgattcc  InF-gRNA-R:
            tccatctttgcattaaaaaaagcaccgactcggtgc  Ta: 69C - ext.: 2242 bp ->
            51" Screening by colony-qPCR: qPCR-gRNA-F + qPCR-EF1a-R
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..1269
                     /label="sanger (hU6-f)"
                     /ApEinfo_revcolor="#b4abac"
                     /ApEinfo_fwdcolor="#b4abac"
     misc_feature    405..2284
                     /label="filler"
                     /ApEinfo_revcolor="#c6c9d1"
                     /ApEinfo_fwdcolor="#c6c9d1"
     misc_feature    complement(1346..2389)
                     /label="sanger (qPCR-gRNA-R)"
                     /ApEinfo_revcolor="#84b0dc"
                     /ApEinfo_fwdcolor="#84b0dc"
     misc_RNA        2285..2360
                     /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          2335..2353
                     /label="qPCR-gRNA-F"
                     /note="sequence: ttgaaaaagtggcaccgag"
                     /ApEinfo_revcolor="#b7e6d7"
                     /ApEinfo_fwdcolor="#b7e6d7"
     primer          complement(2346..2381)
                     /label="InF-gRNA-R"
                     /note="sequence: tccatctttgcattaaaaaaagcaccgactcggtgc"
                     /ApEinfo_revcolor="#b1ff67"
                     /ApEinfo_fwdcolor="#b1ff67"
     misc_feature    2374..3440
                     /label="sanger (qPCR-gRNA-F)"
                     /ApEinfo_revcolor="#c6c9d1"
                     /ApEinfo_fwdcolor="#c6c9d1"
     CDS             2377..2443
                     /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             2377..2443
                     /ApEinfo_revcolor="#84b0dc"
                     /ApEinfo_fwdcolor="#84b0dc"
                     /translation="MDKVLNREESLQLMDLLGLERS"
     primer          complement(2428..2447)
                     /label="qPCR-EF1a-R"
                     /note="sequence: ttcccactcctttcaagacc"
                     /ApEinfo_revcolor="#75c6a9"
                     /ApEinfo_fwdcolor="#75c6a9"
     promoter        2450..3628
                     /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             3642..4361
                     /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             3642..4361
                     /ApEinfo_revcolor="#84b0dc"
                     /ApEinfo_fwdcolor="#84b0dc"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDGNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKVNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLEFVTAAGITLGMDELYK*"
     promoter        complement(4429..4447)
                     /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    4473..4697
                     /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   4716..4856
                     /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(4861..4912)
                     /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      4931..5311
                     /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(5387..5436)
                     /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(5538..5572)
                     /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(5538..5572)
                     /ApEinfo_revcolor="#84b0dc"
                     /ApEinfo_fwdcolor="#84b0dc"
                     /translation="SRVSVMTVKTS"
     promoter        5668..5772
                     /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             5773..6633
                     /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             5773..6633
                     /ApEinfo_revcolor="#84b0dc"
                     /ApEinfo_fwdcolor="#84b0dc"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYIELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYSPVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRWEPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSALPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGASLIKHW*"
     rep_origin      6804..7392
                     /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 accggagacg gttgtaaatg
      421 agcacacaaa atacacatgc taaaatatta tattctatga cctttataaa atcaaccaaa
      481 atcttctttt taataacttt agtatcaata attagaattt ttatgttcct ttttgcaaac
      541 ttttaataaa aatgagcaaa ataaaaaaac gctagtttta gtaactcgcg ttgttttctt
      601 cacctttaat aatagctact ccaccacttg ttcctaagcg gtcagctcct gcttcaatca
      661 ttttttgagc atcttcaaat gttctaactc caccagctgc tttaactaaa gcattgtctt
      721 taacaactga cttcattagt ttaacatctt caaatgttgc acctgatttt gaaaatcctg
      781 ttgatgtttt aacaaattct aatccagctt caacagctat ttcacaagct ttcatgattt
      841 cttcttttgt taataaacaa ttttccataa tacatttaac aacatgtgat ccagctgctt
      901 tttttacagc tttcatgtct tctaaaacta attcataatt tttgtctttt aatgcaccaa
      961 tatttaatac catatcaatt tctgttgcac catctttaat tgcttcagaa acttcgaatg
     1021 cttttgtagc tgttgtgcat gcacctagag gaaaacctac aacatttgtt attcctacat
     1081 ttgtgccttt taataattct ttacaatagc ttgttcaata tgaattaaca caaactgttg
     1141 caaaatcaaa ttcaattgct tcatcacata attgtttaat ttcagctttc gtagcatctt
     1201 gttttaataa tgtgtgatct atatatttgt ttagtttcat tttttctcct atatattcat
     1261 ttttaatttt aattctttaa taatttcgtc tactttaact ttagcgtttt gaacagattc
     1321 accaacacct ataaaataaa tttttagttt aggttcagtt ccacttgggc gaacagcaaa
     1381 tcatgactta tcttctaaat aaaattttag taagtcttgt cctggcatat tatacattcc
     1441 atcgatgtag tcttcaacat taacaacttt aagtccagca atttgagtta agggtgttgc
     1501 tctcaatgat ttcattaatg gttcaatttt taatttcttt tcttctggtt taaaattcaa
     1561 gtttaaagtg aaagtgtaat atgcacccat ttctttaaat aaatcttcta aatagtctac
     1621 taatgtttta ttttgttttt tataaaatca agcagcctct gctattaata tagaagcttg
     1681 tattccatct ttatctctag ctgagtcatc aattacatat ccataacttt cttcataagc
     1741 aaaaacaaaa tttaatccgt tatcttcttc tttagcaatt tctctaccca ttcatttaaa
     1801 tccagttaaa gtttttacaa tattaactcc atatttttca tgagcgattc tatcacccaa
     1861 atcacttgtt acaaaacttg aatatagagc cggatttttt ggaatgctat ttaagcgttt
     1921 tagatttgat aattttcaat caattaaaat tggtcctgtt tgatttccat ctaatcttac
     1981 aaaatgacca tcatgtttta ttgccattcc aaatctgtca gcatctgggt cattcataat
     2041 aataatatct gcatcatgtt taataccata ttcaagcggt atttttcatg caggatcaaa
     2101 ttctggattt ggatttacaa catttttaaa tgtttcatct tcaaatgcat gctcttcaac
     2161 ctcaataacg ttatatcctg attcacgtaa tatttttggg gtaaatttag ttcctgttcc
     2221 attaactgcg ctaaaaataa tttttaaatc ttttttagct tcttgctctt ttttgtacgt
     2281 ctctgtttta gagctagaaa tagcaagtta aaataaggct agtccgttat caacttgaaa
     2341 aagtggcacc gagtcggtgc tttttttaat gcaaagatgg ataaagtttt aaacagagag
     2401 gaatctttgc agctaatgga ccttctaggt cttgaaagga gtgggaattg gctccggtgc
     2461 ccgtcagtgg gcagagcgca catcgcccac agtccccgag aagttggggg gaggggtcgg
     2521 caattgaacc ggtgcctaga gaaggtggcg cggggtaaac tgggaaagtg atgtcgtgta
     2581 ctggctccgc ctttttcccg agggtggggg agaaccgtat ataagtgcag tagtcgccgt
     2641 gaacgttctt tttcgcaacg ggtttgccgc cagaacacag gtaagtgccg tgtgtggttc
     2701 ccgcgggcct ggcctcttta cgggttatgg cccttgcgtg ccttgaatta cttccacctg
     2761 gctgcagtac gtgattcttg atcccgagct tcgggttgga agtgggtggg agagttcgag
     2821 gccttgcgct taaggagccc cttcgcctcg tgcttgagtt gaggcctggc ctgggcgctg
     2881 gggccgccgc gtgcgaatct ggtggcacct tcgcgcctgt ctcgctgctt tcgataagtc
     2941 tctagccatt taaaattttt gatgacctgc tgcgacgctt tttttctggc aagatagtct
     3001 tgtaaatgcg ggccaagatc tgcacactgg tatttcggtt tttggggccg cgggcggcga
     3061 cggggcccgt gcgtcccagc gcacatgttc ggcgaggcgg ggcctgcgag cgcggccacc
     3121 gagaatcgga cgggggtagt ctcaagctgg ccggcctgct ctggtgcctg gcctcgcgcc
     3181 gccgtgtatc gccccgccct gggcggcaag gctggcccgg tcggcaccag ttgcgtgagc
     3241 ggaaagatgg ccgcttcccg gccctgctgc agggagctca aaatggagga cgcggcgctc
     3301 gggagagcgg gcgggtgagt cacccacaca aaggaaaagg gcctttccgt cctcagccgt
     3361 cgcttcatgt gactccacgg agtaccgggc gccgtccagg cacctcgatt agttctcgag
     3421 cttttggagt acgtcgtctt taggttgggg ggaggggttt tatgcgatgg agtttcccca
     3481 cactgagtgg gtggagactg aagttaggcc agcttggcac ttgatgtaat tctccttgga
     3541 atttgccctt tttgagtttg gatcttggtt cattctcaag cctcagacag tggttcaaag
     3601 tttttttctt ccatttcagg tgtcgtgagc ggccgctcga gatggtgagc aagggcgagg
     3661 agctgttcac cggggtggtg cccatcctgg tcgagctgga cggcgacgta aacggccaca
     3721 agttcagcgt gtccggcgag ggcgagggcg atgccaccta cggcaagctg accctgaagt
     3781 tcatctgcac caccggcaag ctgcccgtgc cctggcccac cctcgtgacc accctgacct
     3841 acggcgtgca gtgcttcagc cgctaccccg accacatgaa gcagcacgac ttcttcaagt
     3901 ccgccatgcc cgaaggctac gtccaggagc gcaccatctt cttcaaggac gacggcaact
     3961 acaagacccg cgccgaggtg aagttcgagg gcgacaccct ggtgaaccgc atcgagctga
     4021 agggcatcga cttcaaggag gacggcaaca tcctggggca caagctggag tacaactaca
     4081 acagccacaa cgtctatatc atggccgaca agcagaagaa cggcatcaag gtgaacttca
     4141 agatccgcca caacatcgag gacggcagcg tgcagctcgc cgaccactac cagcagaaca
     4201 cccccatcgg cgacggcccc gtgctgctgc ccgacaacca ctacctgagc acccagtccg
     4261 ccctgagcaa agaccccaac gagaagcgcg atcacatggt cctgctggag ttcgtgaccg
     4321 ccgccgggat cactctcggc atggacgagc tgtacaagta agagcgcacc cgtccgagct
     4381 aaggcgcgcc atacatgcat ggtgtacaag taatctagag ggccctattc tatagtgtca
     4441 cctaaatgct agagctcgct gatcagcctc gactgtgcct tctagttgcc agccatctgt
     4501 tgtttgcccc tcccccgtgc cttccttgac cctggaaggt gccactccca ctgtcctttc
     4561 ctaataaaat gaggaaattg catcgcattg tctgagtagg tgtcattcta ttctgggggg
     4621 tggggtgggg caggacagca agggggagga ttgggaagac aatagcaggc atgctgggga
     4681 tgcggtgggc tctatggctt ctgaggcggg gccgcaggaa cccctagtga tggagttggc
     4741 cactccctct ctgcgcgctc gctcgctcac tgaggccggg cgaccaaagg tcgcccgacg
     4801 cccgggcttt gcccgggcgg cctcagtgag cgagcgagcg cgcagctgcc tgcaggggcg
     4861 cctgatgcgg tattttctcc ttacgcatct gtgcggtatt tcacaccgca tacgtcaaag
     4921 caaccatagt acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc
     4981 agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc
     5041 tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg
     5101 ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgatttggg tgatggttca
     5161 cgtagtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc
     5221 tttaatagtg gactcttgtt ccaaactgga acaacactca accctatctc gggctattct
     5281 tttgatttat aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa
     5341 caaaaattta acgcgaattt taacaaaata ttaacgttta caattttatg gtgcactctc
     5401 agtacaatct gctctgatgc cgcatagtta agccagcccc gacacccgcc aacacccgct
     5461 gacgcgccct gacgggcttg tctgctcccg gcatccgctt acagacaagc tgtgaccttc
     5521 tccgggagct gcatgtgtca gaggttttca ccgtcatcac cgaaacgcgc gatacgaaag
     5581 ggcctcgtga tacgcctatt tttataggtt aatgtcatga taataatggt ttcttagacg
     5641 tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata
     5701 cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga
     5761 aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca
     5821 ttttgccttc ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat
     5881 cagttgggtg cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag
     5941 agttttcgcc ccgaagaacg ttttccaatg atgagcactt ttaaagttct gctatgtggc
     6001 gcggtattat cccgtattga cgccgggcaa gagcaactcg gtcgccgcat acactattct
     6061 cagaatgact tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca
     6121 gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt
     6181 ctgacaacga tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat
     6241 gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt
     6301 gacaccacga tgcctgtagc aatggcaaca acgttgcgca aactattaac tggcgaacta
     6361 cttactctag cttcccggca acaattaata gactggatgg aggcggataa agttgcagga
     6421 ccacttctgc gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt
     6481 gagcgtgggt ctcgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc
     6541 gtagttatct acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct
     6601 gagataggtg cctcactgat taagcattgg taactgtcag accaagttta ctcatatata
     6661 ctttagattg atttaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttttt
     6721 gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc
     6781 gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg
     6841 caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact
     6901 ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt ccttctagtg
     6961 tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg
     7021 ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac
     7081 tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca
     7141 cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga
     7201 gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc
     7261 ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct
     7321 gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg
     7381 agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct
     7441 tttgctcaca tgt
//