LOCUS       sequence-310161-        7205 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For transient expression of mouse RAD21 (SCC1) tagged with mKate2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7205)
  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 7205)
  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: pKS035 - pCAGGS-3XFLAG-Rad21-mKate2-bpA
FEATURES             Location/Qualifiers
     source          1..7205
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        5..384
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     intron          662..1678
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1602..1624
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1686..1705
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     CDS             1765..1830
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             3778..4473
                     /codon_start=1
                     /product="monomeric far-red fluorescent protein (Shcherbo 
                     et al., 2009)"
                     /label=mKate2
                     /note="mammalian codon-optimized"
                     /translation="MVSELIKENMHMKLYMEGTVNNHHFKCTSEGEGKPYEGTQTMRIK
                     AVEGGPLPFAFDILATSFMYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLT
                     ATQDTSLQDGCLIYNVKIRGVNFPSNGPVMQKKTLGWEASTETLYPADGGLEGRADMAL
                     KLVGGGHLICNLKTTYRSKKPAKNLKMPGVYYVDRRLERIKEADKETYVEQHEVAVARY
                     CDLPSKLGHR"
     primer_bind     complement(4564..4583)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    4629..4684
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(4683..4702)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(5045..5061)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5045..5061)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5058..5080)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5069..5085
                     /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(5093..5099,5100..5117,5118..5123))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5138..5159
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5284..5301)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5455..6043)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5535..5554)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6214..7005,7006..7074))
                     /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     6837..6856
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7075..7179)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ggtcgacatt gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat
       61 agcccatata tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg
      121 cccaacgacc cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata
      181 gggactttcc attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta
      241 catcaagtgt atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc
      301 gcctggcatt atgcccagta catgacctta tgggactttc ctacttggca gtacatctac
      361 gtattagtca tcgctattac catggtcgag gtgagcccca cgttctgctt cactctcccc
      421 atctcccccc cctccccacc cccaattttg tatttattta ttttttaatt attttgtgca
      481 gcgatggggg cggggggggg gggggsgcgc gccrggsggg gsggggsggg gsgrggggsg
      541 gggsggggsg aggcggagag gtgcggcggc agccaatcag agcggcgcgc tccgaaagtt
      601 tccttttatg gcgaggcggc ggcggcggcg gccctataaa aagcgaagcg cgcggcgggc
      661 gggagtcgct gcgcgctgcc ttcgccccgt gccccgctcc gccgccgcct cgcgccgccc
      721 gccccggctc tgactgaccg cgttactccc acaggtgagc gggcgggacg gcccttctcc
      781 tccgggctgt aattagcgct tggtttaatg acggcttgtt tcttttctgt ggctgcgtga
      841 aagccttgag gggctccggg agggcccttt gtgcgggggg agcggctcgg ggggtgcgtg
      901 cgtgtgtgtg tgcgtgggga gcgccgcgtg cggctccgcg ctgcccggcg gctgtgagcg
      961 ctgcgggcgc ggcgcggggc tttgtgcgct ccgcagtgtg cgcgagggga gcgcggccgg
     1021 gggcggtgcc ccgcggtgcg gggggggctg cgaggggaac aaaggctgcg tgcggggtgt
     1081 gtgcgtgggg gggtgagcag ggggtgtggg cgcgtcggtc gggctgcaac cccccctgca
     1141 cccccctccc cgagttgctg agcacggccc ggcttcgggt gcggggctcc gtacggggcg
     1201 tggcgcgggg ctcgccgtgc cgggcggggg gtggcggcag gtgggggtgc cgggcggggc
     1261 ggggccgcct cgggccgggg agggctcggg ggaggggcgc ggcggccccc ggagcgccgg
     1321 cggctgtcga ggcgcggcga gccgcagcca ttgcctttta tggtaatcgt gcgagagggc
     1381 gcagggactt cctttgtccc aaatctgtgc ggagccgaaa tctgggaggc gccgccgcac
     1441 cccctctagc gggcgcgggg cgaagcggtg cggcgccggc aggaaggaaa tgggcgggga
     1501 gggccttcgt gcgtcgccgc gccgccgtcc ccttctccct ctccagcctc ggggctgtcc
     1561 gcggggggac ggctgccttc gggggggacg gggcagggcg gggttcggct tctggcgtgt
     1621 gaccggcggc tctagagcct ctgctaacca tgttcatgcc ttcttctttt tcctacagct
     1681 cctgggcaac gtgctggtta ttgtgctgtc tcatcatttt ggcaaagaat tcctcgagaa
     1741 ttctcacgcg tgaattccac catggactat aaggaccacg acggagacta caaggatcat
     1801 gatattgatt acaaagacga tgacgataag accggtgggg gagcaggagg catgttctac
     1861 gcacattttg tcctcagtaa acgagggcct ctggccaaaa tctggctggc ggcccattgg
     1921 gacaagaagc taaccaaagc ccatgtattt gagtgcaact tagagagcag tgtggagagc
     1981 atcatctcac caaaggtgaa gatggcgctg cggacgtcag gacaccttct cctgggagta
     2041 gtccgcatct atcacaggaa agccaaatac ctcctcgcag actgtaatga agcatttatt
     2101 aaaataaaga tggcgtttcg gccaggtgtt gtcgatctac ctgaggaaaa tcgggaagca
     2161 gcttataatg ccattacttt acctgaggaa ttccacgatt ttgatcagcc actgccagat
     2221 ttagatgata ttgacgtcgc ccagcagttc agcctgaacc aaagcagagt agaagagata
     2281 accatgagag aagaagtcgg aaacatcagt atcctacagg aaaatgactt tggtgacttc
     2341 ggaatggatg accgtgaaat aatgagagaa ggcagtgctt tcgaggatga cgacatgtta
     2401 gtgagcacca gcgcttccaa ccttctcctc gagccagagc agagcaccag caacctgaat
     2461 gaaaagatga atcacttaga gtacgaagac cagtacaaag atgacaattt tggagaagga
     2521 aatgatggcg gtatattaga tgacaaactt ataagtaata atgatggtgg catctttgac
     2581 gatccccctg ccttgtctga ggcaggggtc atgttgccag agcaacctgc acatgatgac
     2641 atggatgaag atgacaatgg ctcactgggt gggccggata gtcccgactc tgtggatcct
     2701 gtcgaaccga tgccaactat gactgatcag acaactctcg tcccaaacga ggaagaagct
     2761 tttgcgttgg agcccattga tataactgtc aaagagacaa aagccaagag gaagaggaag
     2821 ctgattgttg acagtgtcaa agaattggat agtaagacca ttagagccca gcttagcgat
     2881 tattctgata ttgttacgac tctggacctg gctccgccaa ccaagaagct tatgatgtgg
     2941 aaagagacag gaggagtgga aaagctcttc tccttaccag cacagcccct gtggaataac
     3001 cggctactga agctcttcac acgctgcctt accccacttg taccagaaga ccttaggaag
     3061 agaaggaaag ggggagaggc agataatctg gatgagttcc tcaaagagtt tgagaatcca
     3121 gaggttccca gagaggagca gcagccacag cagcagcagc cacagccgca gcgagatgtc
     3181 atcgatgagc ccattataga agagccaagc cgcctccagg actcagtgat ggaggccagc
     3241 agaacaacca tagaagaatc agccatgccc ccaccacccc ctcaaggagt taagcggaaa
     3301 gccgggcaaa tagacccaga gccttcgata cctcctcagc aggtagagca aatggaaata
     3361 ccaccagtag aacttccccc agaggagcct ccaaatatct gtcagctgat cccggagtta
     3421 gagctcctac cggagaagga gaaggaaaaa gagaaggaga aggaagagga ggaggaggag
     3481 gaggatgaag atgcttcagg gggtgatcag gatcaagagg aaaggagatg gaacaaacgc
     3541 actcagcaga tgcttcatgg tcttcagcga gctcttgcta aaactggagc agagtctatc
     3601 agtttgcttg agctgtgtcg aaacacaaac cgaaagcagg cagcagcaaa gttctacagc
     3661 tttttggttc ttaagaagca gcaagccatc gagctcacac aggaagagcc gtacagtgac
     3721 atcattgcaa cccctggacc acggttccat attatcctta aggggggagc aggaggcatg
     3781 gtgagcgagc tgattaagga gaacatgcac atgaagctgt acatggaggg caccgtgaac
     3841 aaccaccact tcaagtgcac atccgagggc gaaggcaagc cctacgaggg cacccagacc
     3901 atgagaatca aggcggtcga gggcggccct ctccccttcg ccttcgacat cctggctacc
     3961 agcttcatgt acggcagcaa aaccttcatc aaccacaccc agggcatccc cgacttcttt
     4021 aagcagtcct tccccgaggg cttcacatgg gagagagtca ccacatacga agacgggggc
     4081 gtgctgaccg ctacccagga caccagcctc caggacggct gcctcatcta caacgtcaag
     4141 atcagagggg tgaacttccc atccaacggc cctgtgatgc agaagaaaac actcggctgg
     4201 gaggcctcca ccgagaccct gtaccccgct gacggcggcc tggaaggcag agccgacatg
     4261 gccctgaagc tcgtgggcgg gggccacctg atctgcaact tgaagaccac atacagatcc
     4321 aagaaacccg ctaagaacct caagatgccc ggcgtctact atgtggacag aagactggaa
     4381 agaatcaagg aggccgacaa agagacctac gtcgagcagc acgaggtggc tgtggccaga
     4441 tactgcgacc tccctagcaa actggggcac agaaagtcgc gataagctag ctcaggtgca
     4501 ggctgcctat cagaaggtgg tggctggtgt ggccaatgcc ctggctcaca aataccactg
     4561 agatcttttt ccctctgcca aaaattatgg ggacatcatg aagccccttg agcatctgac
     4621 ttctggctaa taaaggaaat ttattttcat tgcaatagtg tgttggaatt ttttgtgtct
     4681 ctcactcgga aggacatatg ggagggcaaa tcatttaaaa catcagaatg agtatttggt
     4741 ttagagtttg gcaacatatg cccatatgct ggctgccatg aacaaaggtt ggctataaag
     4801 aggtcatcag tatatgaaac agccccctgc tgtccattcc ttattccata gaaaagcctt
     4861 gacttgaggt tagatttttt ttatattttg ttttgtgtta tttttttctt taacatccct
     4921 aaaattttcc ttacatgttt tactagccag atttttcctc ctctcctgac tactcccagt
     4981 catagctgtc cctcttctct tatggagatc cctcgacctg cagcccaagc ttggcgtaat
     5041 catggtcata gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacaacatac
     5101 gagccggaag cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa
     5161 ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag cggatccgct
     5221 gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc
     5281 ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca
     5341 ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg
     5401 agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca
     5461 taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa
     5521 cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc
     5581 tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc
     5641 gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct
     5701 gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg
     5761 tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag
     5821 gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta
     5881 cggctacact agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg
     5941 aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt
     6001 tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt
     6061 ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag
     6121 attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat
     6181 ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc
     6241 tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat
     6301 aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc
     6361 acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag
     6421 aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag
     6481 agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt
     6541 ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg
     6601 agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt
     6661 tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc
     6721 tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc
     6781 attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa
     6841 taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg
     6901 aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc
     6961 caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag
     7021 gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt
     7081 cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt
     7141 tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc
     7201 acctg
//
