LOCUS       sequence-310157-        6980 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Targeting vector to introduce an AID-eGFP cassette at the mouse 
            Cdca5 (SORORIN) locus using BLASTICIDIN selection. Auxin-inducible 
            degron system. Designed to be used with sgRNA GGGATGCCCGTCATTAAGTG.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6980)
  AUTHORS   Nora EP, Caccianini L, Fudenberg G, So K, Kameswaran V, Nagle A, 
            Uebersohn A, Hajj B, Saux AL, Coulon A, Mirny LA, Pollard KS, Dahan 
            M, Bruneau BG
  TITLE     Molecular basis of CTCF binding polarity in genome folding.
  JOURNAL   Nat Commun. 2020 Nov 5;11(1):5612. doi: 10.1038/s41467-020-19283-x.
  PUBMED    33154377
REFERENCE   2  (bases 1 to 6980)
  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: "Nat 
            Commun."; date: "2020-11-5"; volume: "11(1):5612. doi"; pages: " 
            10.1038/s41467-020-19283-x"
COMMENT     Sequence Label: pEN487 - Sororin-AID[71-114]-eGFP-FRT-Blast-FR...
FEATURES             Location/Qualifiers
     source          1..6980
                     /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"
     CDS             1515..1646
                     /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"
     CDS             join(1656..1658,1659..2372)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     CDS             join(1656..1658,1659..2369)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(1698..1719)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1959..1978)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2306..2327
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     protein_bind    2386..2419
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     promoter        2483..2982
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(2518..2541)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     2915..2934
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     promoter        2994..3041
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     regulatory      3066..3075
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3072..3470
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     primer_bind     complement(3506..3523)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3512..3736
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     protein_bind    3791..3824
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     primer_bind     complement(4739..4755)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4739..4755)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     promoter        complement(4792..4810)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(4831..4847)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4831..4847)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4844..4866)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4855..4871
                     /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(4879..4885,4886..4903,4904..4909))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4924..4945
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5062..5079)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5233..5821)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5313..5332)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5992..6783,6784..6852))
                     /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     6615..6634
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6853..6957)
                     /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 gtcgacactt ggacaaggtc ctgcactcca gccctacaac atgaagtaca
      721 aagcaacagt agaaactccg tgtgtcctgt aggcagtctt tccatcctga acccctcgct
      781 cacgagactt ctctcctgtc ctgatacaga tctcttttat tttggaaaaa gagaacaacc
      841 ctccgctcaa ggtacccacg aaagaagacc tcttcaagac ttgtagtgtc cctggtactc
      901 cctccagcac ccccgtgctg tacactcaga atgttgagcc tgactctgga gaagcagagc
      961 tggacagcag agacctggaa atgtctcaga aagtcaggcg gtcgtacagt cggctacaga
     1021 gccttggttg tgcctcaacc tccacccctg gccgccgatc cttctttggc tttgagggac
     1081 ctgacgactt gcctggagtc tcacctgtcg tgtgttcaaa gttaatcgag accccaaagg
     1141 tacctgcaaa ggacttggtt ccggctagaa cgaaggactt ggttccagat agtacgaagg
     1201 acttggttcc ggctagaacg ctccctggaa tctctccccc agttgtgaaa gagaaacgaa
     1261 agaagaaggt tcctgaaatc ctggtgagtg gggccagact tctctgactc ttggtgggaa
     1321 ggcaatgtct gaggtgtggg tctagtgctc tggccaacat ggctgctctt ccctcccact
     1381 catgtgccct catttctttc caccagaaat cagagctgga taagtgggct gtggccatga
     1441 atgctgaatt tgaagctgcc gagcagtttg agcttctgat tgaaggggga gcaggaggcg
     1501 tcgacggtat cgatcctaaa gatccagcca aacctccggc caaggcacaa gttgtgggat
     1561 ggccaccggt gagatcatac cggaagaacg tgatggtttc ctgccaaaaa tcaagcggtg
     1621 gcccggaggc ggcggcgttc gtgaagaagc ttgatgtgag caagggcgag gagctgttca
     1681 ccggggtggt gcccatcctg gtcgagctgg acggcgacgt aaacggccac aagttcagcg
     1741 tgtccggcga gggcgagggc gatgccacct acggcaagct gaccctgaag ttcatctgca
     1801 ccaccggcaa gctgcccgtg ccctggccca ccctcgtgac caccctgacc tacggcgtgc
     1861 agtgcttcag ccgctacccc gaccacatga agcagcacga cttcttcaag tccgccatgc
     1921 ccgaaggcta cgtccaggag cgcaccatct tcttcaagga cgacggcaac tacaagaccc
     1981 gcgccgaggt gaagttcgag ggcgacaccc tggtgaaccg catcgagctg aagggcatcg
     2041 acttcaagga ggacggcaac atcctggggc acaagctgga gtacaactac aacagccaca
     2101 acgtctatat catggccgac aagcagaaga acggcatcaa ggtgaacttc aagatccgcc
     2161 acaacatcga ggacggcagc gtgcagctcg ccgaccacta ccagcagaac acccccatcg
     2221 gcgacggccc cgtgctgctg cccgacaacc actacctgag cacccagtcc gccctgagca
     2281 aagaccccaa cgagaagcgc gatcacatgg tcctgctgga gttcgtgacc gccgccggga
     2341 tcactctcgg catggacgag ctgtacaagt aagatatcga attccgaagt tcctattctc
     2401 tagaaagtat aggaacttca ggtctgaaga ggagtttacg tccagccaag ctagcttggc
     2461 tgcaggtcgt cgaaattcta ccgggtaggg gaggcgcttt tcccaaggca gtctggagca
     2521 tgcgctttag cagccccgct gggcacttgg cgctacacaa gtggcctctg gcctcgcaca
     2581 cattccacat ccaccggtag gcgccaaccg gctccgttct ttggtggccc cttcgcgcca
     2641 ccttctactc ctcccctagt caggaagttc ccccccgccc cgcagctcgc gtcgtgcagg
     2701 acgtgacaaa tggaagtagc acgtctcact agtctcgtgc agatggacag caccgctgag
     2761 caatggaagc gggtaggcct ttggggcagc ggccaatagc agctttgctc cttcgctttc
     2821 tgggctcaga ggctgggaag gggtgggtcc gggggcgggc tcaggggcgg gctcaggggc
     2881 ggggcgggcg cccgaaggtc ctccggaggc ccggcattct gcacgcttca aaagcgcacg
     2941 tctgccgcgc tgttctcctc ttcctcatct ccgggccttt cgacctgcag cctgttgaca
     3001 attaatcatc ggcatagtat atcggcatag tataatacga caaggtgagg aactaaaccc
     3061 atatggccac catggccaag cctttgtctc aagaagaatc caccctcatt gaaagagcaa
     3121 cggctacaat caacagcatc cccatctctg aagactacag cgtcgccagc gcagctctct
     3181 ctagcgacgg ccgcatcttc actggtgtca atgtatatca ttttactggg ggaccttgtg
     3241 cagaactcgt ggtgctgggc actgctgctg ctgcggcagc tggcaacctg acttgtatcg
     3301 tcgcgatcgg aaatgagaac aggggcatct tgagcccctg cggacggtgc cgacaggtgc
     3361 ttctcgatct gcatcctggg atcaaagcca tagtgaagga cagtgatgga cagccgacgg
     3421 cagttgggat tcgtgaattg ctgccctctg gttatgtgtg ggagggctaa cctgagggat
     3481 caattctcta gagctcgctg atcagcctcg actgtgcctt ctagttgcca gccatctgtt
     3541 gtttgcccct cccccgtgcc ttccttgacc ctggaaggtg ccactcccac tgtcctttcc
     3601 taataaaatg aggaaattgc atcgcattgt ctgagtaggt gtcattctat tctggggggt
     3661 ggggtggggc aggacagcaa gggggaggat tgggaagaca atagcaggca tgctggggat
     3721 gcggtgggct ctatggcttc tgaggcggaa agaaccagct ggggctcgac tagagcttgc
     3781 ggaacccttc gaagttccta ttctctagaa agtataggaa cttcatcagt caggtacata
     3841 atggatccga tgaactggga tgcccgtcat taacacgcat cacctggctg gctctcccct
     3901 cttcctgtac atattgcctt ttgagtagag caggatatgt ggggtccctt tcactagcct
     3961 tgttacccag aaaggtattg ctggcctctt gttatattgt gtctccttgc cctggactgt
     4021 ctacctctac agcgttgggg atgagaatcc actgctctga accggaactg tcagaaacag
     4081 ggttagccat gggctgtctc tgctgttgca ttcagccacc aaccagcaga gaggttttgt
     4141 gttctgcttt ggttctgtag agttgagaga tctcaaagac cagggagaca gttaaatgct
     4201 tgagggagga tttttacatt gagtctctgg agtccaggat atgaacggga ggaagcataa
     4261 ttggttttta acacagtgac ttatgtagga actggtccgt gtctgtgctc tgctgttgac
     4321 attggtgaac ggggtgctgc tgcttacagc aaagtgggta cacttgatag cgtgggcctc
     4381 ggatgcacca gtgtaagctt tatagtctct gctttgacct ctttcctcgc ccacatctgt
     4441 ttctttgaat acaagctgtg tggcctcagc cctagacacc atgtcacctc atctctcacc
     4501 tactagacag cttttgctgt gagcttaaaa ctattgctct catgtgagac caaccattcc
     4561 tgggggttgc tgggaacaaa agtggatgta tcatggcttc taaggttcct tcctgcctgc
     4621 cacctctcaa tcatggtaaa agctgtggtt taaaggccat tggccttcca acctttacag
     4681 ttcacgagag cagaagctgt ggggaatgag agctcaggta agctcctgct ccaccaggga
     4741 tccactagtt ctagagcggc cgccaccgcg gtggagctcc agcttttgtt ccctttagtg
     4801 agggttaatt tcgagcttgg cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta
     4861 tccgctcaca attccacaca acatacgagc cggaagcata aagtgtaaag cctggggtgc
     4921 ctaatgagtg agctaactca cattaattgc gttgcgctca ctgcccgctt tccagtcggg
     4981 aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg
     5041 tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg
     5101 gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa
     5161 cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc
     5221 gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc
     5281 aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag
     5341 ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct
     5401 cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta
     5461 ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc
     5521 cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc
     5581 agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt
     5641 gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct
     5701 gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc
     5761 tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca
     5821 agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta
     5881 agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa
     5941 atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg
     6001 cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg
     6061 actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc
     6121 aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc
     6181 cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa
     6241 ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc
     6301 cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg
     6361 ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc
     6421 cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat
     6481 ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg
     6541 tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc
     6601 ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg
     6661 aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat
     6721 gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg
     6781 gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg
     6841 ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct
     6901 catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac
     6961 atttccccga aaagtgccac
//
