LOCUS       sequence-492914-        6293 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Donor plasmid for Crispr/Cas9 gene editing of the human AVIL locus.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6293)
  TITLE     deNooij lab plasmids 2025
REFERENCE   2  (bases 1 to 6293)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"
COMMENT     Sequence Label: pDonor AVIL-P2A-iCre
FEATURES             Location/Qualifiers
     source          1..6293
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      1..589
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     490..509
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     polyA_signal    713..794
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(750..769)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     804..823
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             1867..1923
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             1930..1950
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     rep_origin      complement(3912..4367)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4049..4070)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4261..4280
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4394..4498
                     /gene="bla"
                     /label=AmpR promoter
     promoter        4500..4857
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        4500..4691
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(4523..4543)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4708..4843
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4892..5686
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             4892..5686
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4946..4965)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5556..5575
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5874..5893)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5918..5965
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
ORIGIN
        1 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
       61 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
      121 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
      181 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
      241 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
      301 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
      361 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
      421 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
      481 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
      541 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
      601 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
      661 gttatcccct gattctgtgg ataaccgtat taccgccatg cattagttat taaacttgtt
      721 tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc
      781 atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat ctgcacggtg
      841 gctcatacct gtaatcccag cactttggga gggtgaggtg ggtggatcac gaggtcagga
      901 gttcgagact ggcctggcca atatggtgaa accccgtctg tactaaaaat acaaaaatta
      961 gccaggcgtg gtggtgggtg cctgtagtcc cagctacttg ggaggctgag ggaggagaac
     1021 tgcttgaacc cgggaggcag aggttgcagt gagccgagat tgtgccactg tactccagcc
     1081 tgggccacaa agcaagaatc tgtctcaaaa aaaaaaaaaa agaaaagaaa agaataaatt
     1141 tcttttcccc ttgaagaagt tgatttaggc acagactctg gactctggat ttcccacaat
     1201 gtcttatcta gtcaactcaa gtatctggac tacaattttc ttgaaagcaa agcccatata
     1261 ttaataatct ttacttgtat ataaatattc aataaatcat taagtaaatg tgtagaagaa
     1321 ttttatgctc aataagatcc acccgatcat gcatttgaaa atttatgggt aaaaatggta
     1381 gttacgggca gaaaacaagg aaaaccatag ctcaatttag ttcttcattc tgccaaaatt
     1441 tgatgtctgt aattcatcct taatacataa cccatttatt gccttctggc aagaaactgg
     1501 cgcttgctgg cactgtgagc gcagtgaatc cagtaagctt ctgccacagg ccaaaatggg
     1561 actccctgac ctgaagaaca cctctccaat gaataattta aatctttaac agcctttctg
     1621 agaaattcat tctacttccc tcttcaagga tccacctaga actccctctt gctccaactt
     1681 cctcaacttc tgttggcaat gatgtcatgt cttccttcta actagaaact ttgttttctc
     1741 tcttagaatt acctctctga acaggacttt gtgtctgtgt ttggcatcac aagagggcaa
     1801 tttgcagctc tgcctggctg gaaacagctc caaatgaaga aagaaaaggg acttttcgga
     1861 tccggagcca cgaacttctc tctgttaaag caagcaggag acgtggaaga aaaccccggt
     1921 cctatggtgc ccaagaagaa gaggaaagtc tccaacctgc tgactgtgca ccaaaacctg
     1981 cctgccctcc ctgtggatgc cacctctgat gaagtcagga agaacctgat ggacatgttc
     2041 agggacaggc aggccttctc tgaacacacc tggaagatgc tcctgtctgt gtgcagatcc
     2101 tgggctgcct ggtgcaagct gaacaacagg aaatggttcc ctgctgaacc tgaggatgtg
     2161 agggactacc tcctgtacct gcaagccaga ggcctggctg tgaagaccat ccaacagcac
     2221 ctgggccagc tcaacatgct gcacaggaga tctggcctgc ctcgcccttc tgactccaat
     2281 gctgtgtccc tggtgatgag gagaatcaga aaggagaatg tggatgctgg ggagagagcc
     2341 aagcaggccc tggcctttga acgcactgac tttgaccaag tcagatccct gatggagaac
     2401 tctgacagat gccaggacat caggaacctg gccttcctgg gcattgccta caacaccctg
     2461 ctgcgcattg ccgaaattgc cagaatcaga gtgaaggaca tctcccgcac cgatggtggg
     2521 agaatgctga tccacattgg caggaccaag accctggtgt ccacagctgg tgtggagaag
     2581 gccctgtccc tgggggttac caagctggtg gagagatgga tctctgtgtc tggtgtggct
     2641 gatgacccca acaactacct gttctgccgg gtcagaaaga atggtgtggc tgccccttct
     2701 gccacctccc aactgtccac ccgggccctg gaagggatct ttgaggccac ccaccgcctg
     2761 atctatggtg ccaaggatga ctctgggcag agatacctgg cctggtctgg ccactctgcc
     2821 agagtgggtg ctgccaggga catggccagg gctggtgtgt ccatccctga aatcatgcag
     2881 gctggtggct ggaccaatgt gaacattgtg atgaactaca tcagaaacct ggactctgag
     2941 actggggcca tggtgaggct gctcgaggat ggggactaaa gcaagaaggc ctatacctat
     3001 tgcaaggcca cagaaaagag cagatagtgc caatatcagg aaataattta tccaccaatt
     3061 tctgcctgac attcagctac ttaatttaga tataatagag tctgcaaatc acggcatgtt
     3121 ctccattttt tctcatcctt gcattccttg cttgttatat acctaaaatg ttaaccatat
     3181 agtttttggg ttttgtggcc ctctagctaa agcctcagca gaaagcacta aaactgcata
     3241 aatctggaga aatcaaaaga aagagaacca aaaaacaatg cttaaaatgt ttaataactt
     3301 tatgtttaat attataccag gacctacctt tgttttcaat tttaagatga ttatttctaa
     3361 aatctattta gcctgtaaat cattgaaatc atatatgcac tccataggca aaatccaaat
     3421 acccagatct gtaatgtgtc aaagcatttt tcacttttca aataaagata cctataatga
     3481 acaaatggag ttattttcag tattctgggt ctgcttgtta cggtaatttt atgttatact
     3541 tgaagaaatg tttcagtact ggggaagtaa agaggcaatg ttaagtgagc tctctgggta
     3601 actggactac tctaatttct gtttctggca caagaatgtc aaaatctgcc ttacagctgg
     3661 ccaggaaacc tcagtaccca cccacccaca catcatccaa aagagaaata ggggcatgtg
     3721 ttattaattt cattaaatga ggccagaacc agtgtgtcta ccactgcaga gcaccttcaa
     3781 agcatactac agacaatcac aaagaagaga acccacattt tccagacaac tctaggtttg
     3841 aatcaataat cctaaagaaa tccatgtggc acaagtaata taactaggga ggttccgagg
     3901 cagaagggcg taaattgtaa gcgttaatat tttgttaaaa ttcgcgttaa atttttgtta
     3961 aatcagctca ttttttaacc aataggccga aatcggcaaa atcccttata aatcaaaaga
     4021 atagaccgag atagggttga gtgttgttcc agtttggaac aagagtccac tattaaagaa
     4081 cgtggactcc aacgtcaaag ggcgaaaaac cgtctatcag ggcgatggcc cactacgtga
     4141 accatcaccc taatcaagtt ttttggggtc gaggtgccgt aaagcactaa atcggaaccc
     4201 taaagggagc ccccgattta gagcttgacg gggaaagccg gcgaacgtgg cgagaaagga
     4261 agggaagaaa gcgaaaggag cgggcgctag ggcgctggca agtgtagcgg tcacgctgcg
     4321 cgtaaccacc acacccgccg cgcttaatgc gccgctacag ggcgcgtcag gtggcacttt
     4381 tcggggaaat gtgcgcggaa cccctatttg tttatttttc taaatacatt caaatatgta
     4441 tccgctcatg agacaataac cctgataaat gcttcaataa tattgaaaaa ggaagagtcc
     4501 tgaggcggaa agaaccagct gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc
     4561 tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac caggtgtgga
     4621 aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca
     4681 accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag ttccgcccat
     4741 tctccgcccc atggctgact aatttttttt atttatgcag aggccgaggc cgcctcggcc
     4801 tctgagctat tccagaagta gtgaggaggc ttttttggag gcctaggctt ttgcaaagat
     4861 cgatcaagag acaggatgag gatcgtttcg catgattgaa caagatggat tgcacgcagg
     4921 ttctccggcc gcttgggtgg agaggctatt cggctatgac tgggcacaac agacaatcgg
     4981 ctgctctgat gccgccgtgt tccggctgtc agcgcagggg cgcccggttc tttttgtcaa
     5041 gaccgacctg tccggtgccc tgaatgaact gcaagacgag gcagcgcggc tatcgtggct
     5101 ggccacgacg ggcgttcctt gcgcagctgt gctcgacgtt gtcactgaag cgggaaggga
     5161 ctggctgcta ttgggcgaag tgccggggca ggatctcctg tcatctcacc ttgctcctgc
     5221 cgagaaagta tccatcatgg ctgatgcaat gcggcggctg catacgcttg atccggctac
     5281 ctgcccattc gaccaccaag cgaaacatcg catcgagcga gcacgtactc ggatggaagc
     5341 cggtcttgtc gatcaggatg atctggacga agagcatcag gggctcgcgc cagccgaact
     5401 gttcgccagg ctcaaggcga gcatgcccga cggcgaggat ctcgtcgtga cccatggcga
     5461 tgcctgcttg ccgaatatca tggtggaaaa tggccgcttt tctggattca tcgactgtgg
     5521 ccggctgggt gtggcggacc gctatcagga catagcgttg gctacccgtg atattgctga
     5581 agagcttggc ggcgaatggg ctgaccgctt cctcgtgctt tacggtatcg ccgctcccga
     5641 ttcgcagcgc atcgccttct atcgccttct tgacgagttc ttctgagcgg gactctgggg
     5701 ttcgaaatga ccgaccaagc gacgcccaac ctgccatcac gagatttcga ttccaccgcc
     5761 gccttctatg aaaggttggg cttcggaatc gttttccggg acgccggctg gatgatcctc
     5821 cagcgcgggg atctcatgct ggagttcttc gcccacccta gggggaggct aactgaaaca
     5881 cggaaggaga caataccgga aggaacccgc gctatgacgg caataaaaag acagaataaa
     5941 acgcacggtg ttgggtcgtt tgttcataaa cgcggggttc ggtcccaggg ctggcactct
     6001 gtcgataccc caccgagacc ccattggggc caatacgccc gcgtttcttc cttttcccca
     6061 ccccaccccc caagttcggg tgaaggccca gggctcgcag ccaacgtcgg ggcggcaggc
     6121 cctgccatag cctcaggtta ctcatatata ctttagattg atttaaaact tcatttttaa
     6181 tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt
     6241 gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttc
//
