LOCUS       sequence-360143-        4609 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  CreOFF knock-in for PSD95-Halo-FKBP (amino acid position: R721).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4609)
  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 4609)
  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 Dlg4-Halo-FKBP KI
FEATURES             Location/Qualifiers
     source          1..4609
                     /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        409..484
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     CDS             612..1502
                     /codon_start=1
                     /product="modified bacterial dehalogenase that forms 
                     covalent bonds with chloroalkane derivatives"
                     /label=HaloTag(R)
                     /translation="MAEIGTGFPFDPHYVEVLGERMHYVDVGPRDGTPVLFLHGNPTSS
                     YVWRNIIPHVAPTHRCIAPDLIGMGKSDKPDLGYFFDDHVRFMDAFIEALGLEEVVLVI
                     HDWGSALGFHWAKRNPERVKGIAFMEFIHPIPTWDEWPEFARETFQAFRTTDVGRKLII
                     DQNVFIEGTLPMGVVRPLTEVEMDHYREPFLNPVDREPLWRFPNELPIAGEPANIVALV
                     EEYMDWLHQSPVPKLLFWGTPGVLIPPAEAARLAKSLPNCKAVDIGPGLNLLQEDNPDL
                     IGSEIARWLSTLEISG"
     CDS             1518..1838
                     /codon_start=1
                     /product="human FK506-binding protein FKBP12"
                     /label=FKBP
                     /translation="GVQVETISPGDGRTFPKRGQTCVVHYTGMLEDGKKFDSSRDRNKP
                     FKFMLGKQEVIRGWEEGVAQMSVGQRAKLTISPDYAYGATGHPGIIPPHATLVFDVELL
                     KLE"
     repeat_region   1910..2050
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   1910..2039
                     /label=AAV2 ITR
     rep_origin      2125..2580
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2212..2231)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2422..2443
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2597..2616)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2716..2738
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2776..2794)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2862..2966
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2967..3035,3036..3827)
                     /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(3185..3204)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3998..4586
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4487..4506
                     /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 cttgtttgaa tcagagtctc tctcgggcgt tttagagcta
      421 gaaatagcaa gttaaaataa ggctagtccg ttatcaactt gaaaaagtgg caccgagtcg
      481 gtgctttttt gttttagagc tagaaatagc aagttaaaat aaggctagtc cgtttttagc
      541 gcgtgcgcca attctgcaga caaatggctc tagaagctta atcagagtct ctctcgggcg
      601 gggctagcgg aatggcagaa atcggtactg gctttccatt cgacccccat tatgtggaag
      661 tcctgggcga gcgcatgcac tacgtcgatg ttggtccgcg cgatggcacc cctgtgctgt
      721 tcctgcacgg taacccgacc tcctcctacg tgtggcgcaa catcatcccg catgttgcac
      781 cgacccatcg ctgcattgct ccagacctga tcggtatggg caaatccgac aaaccagacc
      841 tgggttattt cttcgacgac cacgtccgct tcatggatgc cttcatcgaa gccctgggtc
      901 tggaagaggt cgtcctggtc attcacgact ggggctccgc tctgggtttc cactgggcca
      961 agcgcaatcc agagcgcgtc aaaggtattg catttatgga gttcatccac cctatcccga
     1021 cctgggacga atggccagaa tttgcccgcg agaccttcca ggccttccgc accaccgacg
     1081 tcggccgcaa gctgatcatc gatcagaacg tttttatcga gggtacgctg ccgatgggtg
     1141 tcgtccgccc gctgactgaa gtcgagatgg accattaccg cgagccgttc ctgaatcctg
     1201 ttgaccgcga gccactgtgg cgcttcccaa acgagctgcc aatcgccggt gagccagcga
     1261 acatcgtcgc gctggtcgaa gaatacatgg actggctgca ccagtcccct gtcccgaagc
     1321 tgctgttctg gggcacccca ggcgttctga tcccaccggc cgaagccgct cgcctggcca
     1381 aaagcctgcc taactgcaag gctgtggaca tcggcccggg tctgaatctg ctgcaagaag
     1441 acaacccgga cctgatcggc agcgagatcg cgcgctggct gtccacgctc gagatttccg
     1501 gcagcggagg cagcgctgga gtgcaggtgg aaaccatctc cccaggagac gggcgcacct
     1561 tccccaagcg cggccagacc tgcgtggtgc actacaccgg gatgcttgaa gatggaaaga
     1621 aatttgattc ctcccgggac agaaacaagc cctttaagtt tatgctaggc aagcaggagg
     1681 tgatccgagg ctgggaagaa ggggttgccc agatgagtgt gggtcagaga gccaaactga
     1741 ctatatctcc agattatgcc tatggtgcca ctgggcaccc aggcatcatc ccaccacatg
     1801 ccactctcgt cttcgatgtg gagcttctaa aactggaagg agtcgacgct aaatcagagt
     1861 ctctctcggg cgggacgcgt taacttacgg taaatggccc ggcggccgca ggaaccccta
     1921 gtgatggagt tggccactcc ctctctgcgc gctcgctcgc tcactgaggc cgggcgacca
     1981 aaggtcgccc gacgcccggg ctttgcccgg gcggcctcag tgagcgagcg agcgcgcagc
     2041 tgcctgcagg ggcgcctgat gcggtatttt ctccttacgc atctgtgcgg tatttcacac
     2101 cgcatacgtc aaagcaacca tagtacgcgc cctgtagcgg cgcattaagc gcggcgggtg
     2161 tggtggttac gcgcagcgtg accgctacac ttgccagcgc cttagcgccc gctcctttcg
     2221 ctttcttccc ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg
     2281 ggctcccttt agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt
     2341 tgggtgatgg ttcacgtagt gggccatcgc cctgatagac ggtttttcgc cctttgacgt
     2401 tggagtccac gttctttaat agtggactct tgttccaaac tggaacaaca ctcaactcta
     2461 tctcgggcta ttcttttgat ttataaggga ttttgccgat ttcggtctat tggttaaaaa
     2521 atgagctgat ttaacaaaaa tttaacgcga attttaacaa aatattaacg tttacaattt
     2581 tatggtgcac tctcagtaca atctgctctg atgccgcata gttaagccag ccccgacacc
     2641 cgccaacacc cgctgacgcg ccctgacggg cttgtctgct cccggcatcc gcttacagac
     2701 aagctgtgac cgtctccggg agctgcatgt gtcagaggtt ttcaccgtca tcaccgaaac
     2761 gcgcgagacg aaagggcctc gtgatacgcc tatttttata ggttaatgtc atgataataa
     2821 tggtttctta gacgtcaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt
     2881 tatttttcta aatacattca aatatgtatc cgctcatgag acaataaccc tgataaatgc
     2941 ttcaataata ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc gcccttattc
     3001 ccttttttgc ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa
     3061 aagatgctga agatcagttg ggtgcacgag tgggttacat cgaactggat ctcaacagcg
     3121 gtaagatcct tgagagtttt cgccccgaag aacgttttcc aatgatgagc acttttaaag
     3181 ttctgctatg tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc
     3241 gcatacacta ttctcagaat gacttggttg agtactcacc agtcacagaa aagcatctta
     3301 cggatggcat gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg
     3361 cggccaactt acttctgaca acgatcggag gaccgaagga gctaaccgct tttttgcaca
     3421 acatggggga tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac
     3481 caaacgacga gcgtgacacc acgatgcctg tagcaatggc aacaacgttg cgcaaactat
     3541 taactggcga actacttact ctagcttccc ggcaacaatt aatagactgg atggaggcgg
     3601 ataaagttgc aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata
     3661 aatctggagc cggtgagcgt ggaagccgcg gtatcattgc agcactgggg ccagatggta
     3721 agccctcccg tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa
     3781 atagacagat cgctgagata ggtgcctcac tgattaagca ttggtaactg tcagaccaag
     3841 tttactcata tatactttag attgatttaa aacttcattt ttaatttaaa aggatctagg
     3901 tgaagatcct ttttgataat ctcatgacca aaatccctta acgtgagttt tcgttccact
     3961 gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg
     4021 taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc
     4081 aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata
     4141 ctgttcttct agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta
     4201 catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc
     4261 ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg
     4321 ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac
     4381 agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg
     4441 taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt
     4501 atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct
     4561 cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttt
//
