LOCUS       sequence-275809-        4869 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mouse N-terminal CTCF CRISPR knock-in donor vector with miniAID 
            degron and HaloTag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4869)
  AUTHORS   Xie L, Dong P, Chen X, Hsieh TS, Banala S, De Marzio M, English BP, 
            Qi Y, Jung SK, Kieffer-Kwon KR, Legant WR, Hansen AS, Schulmann A, 
            Casellas R, Zhang B, Betzig E, Lavis LD, Chang HY, Tjian R, Liu Z
  TITLE     3D ATAC-PALM: super-resolution imaging of the accessible genome.
  JOURNAL   Nat Methods. 2020 Apr;17(4):430-436. doi: 10.1038/s41592-020-0775-2.
            Epub 2020 Mar 16.
  PUBMED    32203384
REFERENCE   2  (bases 1 to 4869)
  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: 
            "10.1038/s41592-020-0775-2"; journalName: "Nat Methods"; date: 
            "2020-04"; volume: "17"; issue: "4"; pages: "430-436"
COMMENT     Sequence Label: pLX-N-CTCF-mAID-Halo-donor
FEATURES             Location/Qualifiers
     source          1..4869
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(29..51)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     151..170
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     364..386
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     378..395
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     379..395
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             915..938
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             948..1838
                     /codon_start=1
                     /product="modified bacterial dehalogenase that forms 
                     covalent bonds with chloroalkane derivatives"
                     /label=HaloTag(R)
                     /translation="MAEIGTGFPFDPHYVEVLGERMHYVDVGPRDGTPVLFLHGNPTSS
                     YVWRNIIPHVAPTHRCIAPDLIGMGKSDKPDLGYFFDDHVRFMDAFIEALGLEEVVLVI
                     HDWGSALGFHWAKRNPERVKGIAFMEFIRPIPTWDEWPEFARETFQAFRTTDVGRKLII
                     DQNVFIEGTLPMGVVRPLTEVEMDHYREPFLNPVDREPLWRFPNELPIAGEPANIVALV
                     EEYMDWLHQSPVPKLLFWGTPGVLIPPAEAARLAKSLPNCKAVDIGPGLNLLQEDNPDL
                     IGSEIARWLSTLEISG"
     CDS             1851..1871
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="EDLYFQS"
     CDS             1920..2051
                     /codon_start=1
                     /product="minimal degron from the auxin-responsive IAA17 
                     protein of Arabidopsis thaliana (Morawska and Ulrich, 
                     2013)"
                     /label=AID*
                     /note="acts as an auxin-inducible degron (AID) in the 
                     presence of TIR1"
                     /translation="PKDPAKPPAKAQVVGWPPVRSYRKNVMVSCQKSSGGPEAAAFVK"
     primer_bind     complement(2648..2664)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2648..2664)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2661..2683)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2672..2688
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(2696..2702,2703..2720,2721..2726))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2741..2762
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2879..2896)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3050..3638)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3130..3149)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3809..4600,4601..4669))
                     /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     4432..4451
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4670..4774)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4842..4860
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca
       61 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
      121 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
      181 accatatgcg gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc atcaggcgcc
      241 attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat
      301 tacgccagct ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta acgccagggt
      361 tttcccagtc acgacgttgt aaaacgacgg ccagtgaatt caaaccaaaa ttaacattgt
      421 agtgtttctc ttagactgaa cttctgcagt atatttaaca cctgctgtca ccagttttct
      481 gttaattttg agtataatct agagtcttga tcagtttcag atttttttgt tttgattggc
      541 aagatttata atataggtgg tgaatttttt tttcttccat caaagaggaa gatatctatt
      601 aaataaccat tgatggatgc ctaattcatt cactaaggtg ggttttttgg ggggtcagga
      661 ggttgagttt taggggtttg gcttggtttt tggttcttag agatagggtt tctctgtgca
      721 ggccaggctg gcctcaaact cagaaatctg cctgcctgtg ggttctaagt gctgggatta
      781 aaggtgtgtg caccactacc acctgccact caaatgattt tagtttaaaa tgtgtgcttt
      841 gatttgagca aaccacatca taaataacat ctgtgttctc cttaataaag gaaggggaga
      901 tggaaactag aatggattac aaggatgacg atgacaagct aggctccatg gcagaaatcg
      961 gtactggctt tccattcgac ccccattatg tggaagtcct gggcgagcgc atgcactacg
     1021 tcgatgttgg tccgcgcgat ggcacccctg tgctgttcct gcacggtaac ccgacctcct
     1081 cctacgtgtg gcgcaacatc atcccgcatg ttgcaccgac ccatcgctgc attgctccag
     1141 acctgatcgg tatgggcaaa tccgacaaac cagacctggg ttatttcttc gacgaccacg
     1201 tccgcttcat ggatgccttc atcgaagccc tgggtctgga agaggtcgtc ctggtcattc
     1261 acgactgggg ctccgctctg ggtttccact gggccaagcg caatccagag cgcgtcaaag
     1321 gtattgcatt tatggagttc atccgcccta tcccgacctg ggacgaatgg ccagaatttg
     1381 cccgcgagac cttccaggcc ttccgcacca ccgacgtcgg ccgcaagctg atcatcgatc
     1441 agaacgtttt tatcgagggt acgctgccga tgggtgtcgt ccgcccgctg actgaagtcg
     1501 agatggacca ttaccgcgag ccgttcctga atcctgttga ccgcgagcca ctgtggcgct
     1561 tcccaaacga gctgccaatc gccggtgagc cagcgaacat cgtcgcgctg gtcgaagaat
     1621 acatggactg gctgcaccag tcccctgtcc cgaagctgct gttctggggc accccaggcg
     1681 ttctgatccc accggccgaa gccgctcgcc tggccaaaag cctgcctaac tgcaaggctg
     1741 tggacatcgg cccgggtctg aatctgctgc aagaagacaa cccggacctg atcggcagcg
     1801 agatcgcgcg ctggctgtcg acgctcgaga tttccggcga gccaaccact gaggatctgt
     1861 actttcagag cgataactcc ggccgtacgg ggggagcagg aggcgtcgac ggtatcgatc
     1921 ctaaagatcc agccaaacct ccggccaagg cacaagttgt gggatggcca ccggtgagat
     1981 cataccggaa gaacgtgatg gtttcctgcc aaaaatcaag cggtggcccg gaggcggcgg
     2041 cgttcgtgaa gaagcttgat ggtggaggtt ccggtgaagc tgtcgaggct atcgttgaag
     2101 agtctgaaac tttcattaaa ggaaaagaaa gaaagacata tcagcgcaga agagagggag
     2161 gacaggaaga agatgcttgc cacctgcccc agaaccagac agatgggggt gaggtggtcc
     2221 aggatgtcaa cagcagtgta cagatggtaa tgatggaaca gctggatcct acccttctcc
     2281 agatgaagac tgaagtcatg gagggtacag tggctccgga agcagaggct gcagtggacg
     2341 atacccagat cataaccttg caggttgtaa atatggagga acagcccatt aacataggag
     2401 agcttcagct tgtccaagta cctgttcctg tgacggtacc tgttgctact acttcagtag
     2461 aagaacttca gggggcttat gagaatgaag tgtctaaaga gggccttgca gaaagtgaac
     2521 cgatgatatg tcacacctta cctttgcctg aaggatttca ggtggtgaaa gtgggggcca
     2581 atggagaagt ggaagatccc gggcccgtcg actgcagagg cctgcatgca agcttggcgt
     2641 aatcatggtc atagctgttt cctgtgtgaa attgttatcc gctcacaatt ccacacaaca
     2701 tacgagccgg aagcataaag tgtaaagcct ggggtgccta atgagtgagc taactcacat
     2761 taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt
     2821 aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct tccgcttcct
     2881 cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa
     2941 aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa
     3001 aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc
     3061 tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga
     3121 caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc
     3181 cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt
     3241 ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct
     3301 gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg
     3361 agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta
     3421 gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct
     3481 acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa
     3541 gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt
     3601 gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta
     3661 cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat
     3721 caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa
     3781 gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct
     3841 cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta
     3901 cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct
     3961 caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg
     4021 gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa
     4081 gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt
     4141 cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta
     4201 catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca
     4261 gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta
     4321 ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct
     4381 gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg
     4441 cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac
     4501 tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact
     4561 gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa
     4621 atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt
     4681 ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat
     4741 gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg
     4801 acgtctaaga aaccattatt atcatgacat taacctataa aaataggcgt atcacgaggc
     4861 cctttcgtc
//
