LOCUS       sequence-466501-        4901 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Donor plasmid for tagging mouse CTCF with mAID-eGFP at the 
            N-terminus. Used with KL061 - PX330-sgRNA006_CTCF sgRNA..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4901)
  AUTHORS   Lucero K, Han S, Huang P, Qiu X, Mazzoni EO, Reinberg D
  TITLE     CTCF-RNA interactions orchestrate cell-specific chromatin loop 
            organization
  JOURNAL   bioRxiv, March 19, 2025
REFERENCE   2  (bases 1 to 4901)
  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"; journalName: "bioRxiv, 
            March 19, 2025"
COMMENT     Sequence Label: KL060 - pBlueScript II SK-mAID-EGFP CTCF donor
FEATURES             Location/Qualifiers
     source          1..4901
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(30..458)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(140..161)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     352..371
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     585..607
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     599..616
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     600..616
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     623..642
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        623..641
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     666..682
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     667..683
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             1195..1398
                     /codon_start=1
                     /product="minimal degron from the auxin-responsive IAA17 
                     protein of Arabidopsis thaliana (Kubota et al., 2013)"
                     /label=mini-AID
                     /note="acts as an auxin-inducible degron (AID) in the 
                     presence of TIR1"
                     /translation="KEKSACPKDPAKPPAKAQVVGWPPVRSYRKNVMVSCQKSSGGPEA
                     AAFVKVSMDGAPYLRKIDLRMYK"
     CDS             join(1405..1407,1408..1410,1411..2121)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(1405..1407,1408..1410,1411..2121)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1450..1471)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1711..1730)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2058..2079
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(2660..2676)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2660..2676)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     promoter        complement(2713..2731)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2752..2768)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2752..2768)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2765..2787)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2776..2792
                     /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(2800..2806,2807..2824,2825..2830))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2845..2866
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2983..3000)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3154..3742)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3234..3253)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3913..4704,4705..4773))
                     /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     4536..4555
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4774..4878)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ctaaattgta agcgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc
       61 attttttaac caataggccg aaatcggcaa aatcccttat aaatcaaaag aatagaccga
      121 gatagggttg agtgttgttc cagtttggaa caagagtcca ctattaaaga acgtggactc
      181 caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg aaccatcacc
      241 ctaatcaagt tttttggggt cgaggtgccg taaagcacta aatcggaacc ctaaagggag
      301 cccccgattt agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa
      361 agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac
      421 cacacccgcc gcgcttaatg cgccgctaca gggcgcgtcc cattcgccat tcaggctgcg
      481 caactgttgg gaagggcgat cggtgcgggc ctcttcgcta ttacgccagc tggcgaaagg
      541 gggatgtgct gcaaggcgat taagttgggt aacgccaggg ttttcccagt cacgacgttg
      601 taaaacgacg gccagtgaat tgtaatacga ctcactatag ggcgaattgg gtaccgggcc
      661 ccccctcgag gtcgacggta tcgataagct tgataaccaa aattaacatt gtagtgtttc
      721 tcttagactg aacttctgca gtatatttaa cacctgctgt caccagtttt ctgttaattt
      781 tgagtataat ctagagtctt gatcagtttc agattttttt gttttgattg gcaagattta
      841 taatataggt ggtgaatttt tttttcttcc atcaaagagg aagatatcta ttaaataacc
      901 attgatggat gcctaattca ttcactaagg tgggtttttt ggggggtcag gaggttgagt
      961 tttaggggtt tggcttggtt tttggttctt agagataggg tttctctgtg caggccaggc
     1021 tggcctcaaa ctcagaaatc tgcctgcctg tgggttctaa gtgctgggat taaaggtgtg
     1081 tgcaccacta ccacctgcca ctcaaatgat tttagtttaa aatgtgtgct ttgatttgag
     1141 caaaccacat cataaataac atctgtgttc tccttaataa aggaagggga gatgaaggag
     1201 aagagtgctt gtcctaaaga tccagccaaa cctccggcca aggcacaagt tgtgggatgg
     1261 ccaccggtga gatcataccg gaagaacgtg atggtttcct gccaaaaatc aagcggtggc
     1321 ccggaggcgg cggcgttcgt gaaggtatca atggacggag caccgtactt gaggaaaatc
     1381 gatttgagga tgtataaagc tagcatggtg agcaagggcg aggagctgtt caccggggtg
     1441 gtgcccatcc tggtcgagct ggacggcgac gtaaacggcc acaagttcag cgtgtccggc
     1501 gagggcgagg gcgatgccac ctacggcaag ctgaccctga agttcatctg caccaccggc
     1561 aagctgcccg tgccctggcc caccctcgtg accaccctga cctacggcgt gcagtgcttc
     1621 agccgctacc ccgaccacat gaagcagcac gacttcttca agtccgccat gcccgaaggc
     1681 tacgtccagg agcgcaccat cttcttcaag gacgacggca actacaagac ccgcgccgag
     1741 gtgaagttcg agggcgacac cctggtgaac cgcatcgagc tgaagggcat cgacttcaag
     1801 gaggacggca acatcctggg gcacaagctg gagtacaact acaacagcca caacgtctat
     1861 atcatggccg acaagcagaa gaacggcatc aaggtgaact tcaagatccg ccacaacatc
     1921 gaggacggca gcgtgcagct cgccgaccac taccagcaga acacccccat cggcgacggc
     1981 cccgtgctgc tgcccgacaa ccactacctg agcacccagt ccgccctgag caaagacccc
     2041 aacgagaagc gcgatcacat ggtcctgctg gagttcgtga ccgccgccgg gatcactctc
     2101 ggcatggacg agctgtacaa gggtgcaggc gcccaagctt cgaattctgc agtcgacgaa
     2161 ggtgaagccg tagaggcgat agtcgaggag tctgaaactt tcattaaagg aaaagaaaga
     2221 aagacttacc agagacgccg ggaagggggc caggaagaag atgcttgcca cctgccccag
     2281 aaccagacag atgggggtga ggtggtccag gatgtcaaca gcagtgtaca gatggtaatg
     2341 atggaacagc tggatcctac ccttctccag atgaagactg aagtcatgga gggtacagtg
     2401 gctccggaag cagaggctgc agtggacgat acccagatca taaccttgca ggttgtaaat
     2461 atggaggaac agcccattaa cataggagag cttcagcttg tccaagtacc tgttcctgtg
     2521 acggtacctg ttgctactac ttcagtagaa gaacttcagg gggcttatga gaatgaagtg
     2581 tctaaagagg gccttgcaga aagtgaaccg atgatatgtc acaccttacc tttgcctgaa
     2641 ggatttcagg tggtgaaagg atccactagt tctagagcgg ccgccaccgc ggtggagctc
     2701 cagcttttgt tccctttagt gagggttaat ttcgagcttg gcgtaatcat ggtcatagct
     2761 gtttcctgtg tgaaattgtt atccgctcac aattccacac aacatacgag ccggaagcat
     2821 aaagtgtaaa gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc
     2881 actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg
     2941 cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct
     3001 gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt
     3061 atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc
     3121 caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga
     3181 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata
     3241 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac
     3301 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg
     3361 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc
     3421 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag
     3481 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt
     3541 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt
     3601 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg
     3661 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac
     3721 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca
     3781 gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac
     3841 ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac
     3901 ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt
     3961 tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt
     4021 accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt
     4081 atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc
     4141 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa
     4201 tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg
     4261 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt
     4321 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc
     4381 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt
     4441 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg
     4501 gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac
     4561 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc
     4621 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt
     4681 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg
     4741 aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat attattgaag
     4801 catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa
     4861 acaaataggg gttccgcgca catttccccg aaaagtgcca c
//
