LOCUS       sequence-360208-        3345 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  CreOFF knock-in vector.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3345)
  AUTHORS   Droogers WJ, Willems J, MacGillavry HD, de Jong APH
  TITLE     Duplex Labeling and Manipulation of Neuronal Proteins Using 
            Sequential CRISPR/Cas9 Gene Editing.
  JOURNAL   eNeuro. 2022 Jul 18;9(4):ENEURO.0056-22.2022. doi: 
            10.1523/ENEURO.0056-22.2022.
  PUBMED    35851300
REFERENCE   2  (bases 1 to 3345)
  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"; doi: 
            "10.1523/ENEURO.0056-22"; journalName: "eNeuro"; date: "2022-07-18- 
            18"; volume: "9"; issue: "4"; pages: "ENEURO.0056-22.2022"
COMMENT     Sequence Label: pOC2 cloning template vector
FEATURES             Location/Qualifiers
     source          1..3345
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    complement(39..72)
                     /label=loxP511
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATAC) (Shaw et al., 2021). loxP511 sites are 
                     compatible with each other, but incompatible with wild-type
                     loxP sites."
     protein_bind    complement(343..376)
                     /label=loxP511
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATAC) (Shaw et al., 2021). loxP511 sites are 
                     compatible with each other, but incompatible with wild-type
                     loxP sites."
     misc_RNA        410..485
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     repeat_region   646..786
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   646..775
                     /label=AAV2 ITR
     rep_origin      861..1316
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(948..967)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1158..1179
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(1333..1352)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     1452..1474
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(1512..1530)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1598..1702
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1703..1771,1772..2563)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(1921..1940)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2734..3322
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3223..3242
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 acggttcctg gccttttgct ggccttttgc tcacatgtat aacttcgtat agtataaatt
       61 atacgaagtt atgatccgac gccgccatct ctaggcccgc gccggccccc tcgcacagac
      121 ttgtgggaga agctcggcta ctcccctgcc ccggttaatt tgcatataat atttcctagt
      181 aactatagag gcttaatgtg cgataaaaga cagataatct gttcttttta atactagcta
      241 cattttacat gataggcttg gatttctata agagatacaa atactaaatt attattttaa
      301 aaaacagcac actcgagaac tcaccctaac tgtaaagtta gtataacttc gtatagtata
      361 aattatacga agttataagc cttgtttgtc ttcgtgtaca agaagacctg ttttagagct
      421 agaaatagca agttaaaata aggctagtcc gttatcaact tgaaaaagtg gcaccgagtc
      481 ggtgcttttt tgttttagag ctagaaatag caagttaaaa taaggctagt ccgtttttag
      541 cgcgtgcgcc aattctgcag acaaatggct ctagaagctt gctagcaatt gtcgactcga
      601 ggatcctagg acgcgttaac ttacggtaaa tggcccggcg gccgcaggaa cccctagtga
      661 tggagttggc cactccctct ctgcgcgctc gctcgctcac tgaggccggg cgaccaaagg
      721 tcgcccgacg cccgggcttt gcccgggcgg cctcagtgag cgagcgagcg cgcagctgcc
      781 tgcaggggcg cctgatgcgg tattttctcc ttacgcatct gtgcggtatt tcacaccgca
      841 tacgtcaaag caaccatagt acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt
      901 ggttacgcgc agcgtgaccg ctacacttgc cagcgcctta gcgcccgctc ctttcgcttt
      961 cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcgggggct
     1021 ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgatttggg
     1081 tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga
     1141 gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca actctatctc
     1201 gggctattct tttgatttat aagggatttt gccgatttcg gtctattggt taaaaaatga
     1261 gctgatttaa caaaaattta acgcgaattt taacaaaata ttaacgttta caattttatg
     1321 gtgcactctc agtacaatct gctctgatgc cgcatagtta agccagcccc gacacccgcc
     1381 aacacccgct gacgcgccct gacgggcttg tctgctcccg gcatccgctt acagacaagc
     1441 tgtgaccgtc tccgggagct gcatgtgtca gaggttttca ccgtcatcac cgaaacgcgc
     1501 gagacgaaag ggcctcgtga tacgcctatt tttataggtt aatgtcatga taataatggt
     1561 ttcttagacg tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt
     1621 tttctaaata cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca
     1681 ataatattga aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc ttattccctt
     1741 ttttgcggca ttttgccttc ctgtttttgc tcacccagaa acgctggtga aagtaaaaga
     1801 tgctgaagat cagttgggtg cacgagtggg ttacatcgaa ctggatctca acagcggtaa
     1861 gatccttgag agttttcgcc ccgaagaacg ttttccaatg atgagcactt ttaaagttct
     1921 gctatgtggc gcggtattat cccgtattga cgccgggcaa gagcaactcg gtcgccgcat
     1981 acactattct cagaatgact tggttgagta ctcaccagtc acagaaaagc atcttacgga
     2041 tggcatgaca gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc
     2101 caacttactt ctgacaacga tcggaggacc gaaggagcta accgcttttt tgcacaacat
     2161 gggggatcat gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa
     2221 cgacgagcgt gacaccacga tgcctgtagc aatggcaaca acgttgcgca aactattaac
     2281 tggcgaacta cttactctag cttcccggca acaattaata gactggatgg aggcggataa
     2341 agttgcagga ccacttctgc gctcggccct tccggctggc tggtttattg ctgataaatc
     2401 tggagccggt gagcgtggaa gccgcggtat cattgcagca ctggggccag atggtaagcc
     2461 ctcccgtatc gtagttatct acacgacggg gagtcaggca actatggatg aacgaaatag
     2521 acagatcgct gagataggtg cctcactgat taagcattgg taactgtcag accaagttta
     2581 ctcatatata ctttagattg atttaaaact tcatttttaa tttaaaagga tctaggtgaa
     2641 gatccttttt gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc
     2701 gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat
     2761 ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga
     2821 gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt
     2881 tcttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata
     2941 cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac
     3001 cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg
     3061 ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg
     3121 tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag
     3181 cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct
     3241 ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc
     3301 aggggggcgg agcctatgga aaaacgccag caacgcggcc ttttt
//
