LOCUS       sequence-364584-        5504 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For knocking in FKBP F36V P2A mCerulean.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5504)
  AUTHORS   Asimi V, Sampath Kumar A, Niskanen H, Riemenschneider C, Hetzel S, 
            Naderi J, Fasching N, Popitsch N, Du M, Kretzmer H, Smith ZD, 
            Weigert R, Walther M, Mamde S, Meierhofer D, Wittler L, Buschow R, 
            Timmermann B, Cisse II, Ameres SL, Meissner A, Hnisz D
  TITLE     Hijacking of transcriptional condensates by endogenous retroviruses.
  JOURNAL   Nat Genet. 2022 Aug;54(8):1238-1247. doi: 
            10.1038/s41588-022-01132-w. Epub 2022 Jul 21.
  PUBMED    35864192
REFERENCE   2  (bases 1 to 5504)
  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/s41588-022-01132-w"; journalName: "Nat Genet"; date: 
            "2022-08"; volume: "54"; issue: "8"; pages: "1238-1247"
COMMENT     Sequence Label: pUC19.TRIM28.FKBP.knock-in.mCerulean
FEATURES             Location/Qualifiers
     source          1..5504
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(19..38)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     366..387
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             439..495
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             496..522
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /note="This is a variant of the standard HA tag, which has 
                     sequence YPYDVPDYA. Some anti-HA antibodies may not 
                     recognize this variant."
                     /translation="YPYDVPDYA"
     CDS             496..522
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             526..552
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             562..882
                     /codon_start=1
                     /product="F36V mutant of FK506-binding protein FKBP12"
                     /label=FKBP-V
                     /note="binds synthetic ligands that do not bind wild-type 
                     FKBP"
                     /translation="GVQVETISPGDGRTFPKRGQTCVVHYTGMLEDGKKVDSSRDRNKP
                     FKFMLGKQEVIRGWEEGVAQMSVGQRAKLTISPDYAYGATGHPGIIPPHATLVFDVELL
                     KLE"
     primer_bind     complement(1748..1765)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1748..1764)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1757..1779)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(1973..1992)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2092..2114
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2152..2170)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2238..2342
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2343..2411,2412..3203)
                     /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(2561..2580)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3374..3962
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3863..3882
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4116..4133
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    4250..4271
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(4286..4291,4292..4309,4310..4316)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4324..4340
                     /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)."
     primer_bind     4329..4351
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     4348..4364
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4348..4364
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             join(5217..5219,5220..5222,5223..5504,1..429)
                     /codon_start=1
                     /product="enhanced monomeric variant of CFP (Rizzo et al., 
                     2004)"
                     /label=mCerulean
                     /note=" mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHRFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTWGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNAISDNVYITADKQKNGIK
                     AHFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(5262..5283)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
ORIGIN
        1 cgtaccatct tcttcaagga cgacggcaac tacaagaccc gcgccgaggt gaagttcgag
       61 ggcgacaccc tggtgaaccg catcgagctg aagggcatcg acttcaagga ggacggcaac
      121 atcctggggc acaagctgga gtacaacgcc atcagcgaca acgtctatat caccgccgac
      181 aagcagaaga acggcatcaa ggcccacttc aagatccgcc acaacatcga ggacggcagc
      241 gtgcagctcg ccgaccacta ccagcagaac acccccatcg gcgacggccc cgtgctgctg
      301 cccgacaacc actacctgag cacccagtcc aagctgagca aagaccccaa cgagaagcgc
      361 gatcacatgg tcctgctgga gttcgtgacc gccgccggga tcactctcgg catggacgag
      421 ctgtacaagg gaagcggagc tactaacttc agcctgctga agcaggctgg agacgtggag
      481 gagaaccctg gaccttaccc ctacgacgtg cccgactacg ccggctatcc gtatgatgtc
      541 ccggactatg caggaagcgg aggagtgcag gtggaaacca tctccccagg agacgggcgc
      601 accttcccca agcgcggcca gacctgcgtg gtgcactaca ccgggatgct tgaagatgga
      661 aagaaagttg attcctcccg ggacagaaac aagcccttta agtttatgct aggcaagcag
      721 gaggtgatcc gaggctggga agaaggggtt gcccagatga gtgtgggtca gagagccaaa
      781 ctgactatat ctccagatta tgcctatggt gccactgggc acccaggcat catcccacca
      841 catgccactc tcgtcttcga tgtggagctt ctaaaactgg aaggtggcgg tggctcgggc
      901 ggtggtgggt cgggtggcgg cggatctgcc tcggcggcag cgactgcagc ggcctcggcc
      961 gcgacggccg cctcggcggc ctctggtagc ccagggtcgg gcgagggctc ggcgggcggt
     1021 gagaagcgtc cggctgcttc ctcagccgcg gcggcctctg cagccgcgtc gtcccctgcg
     1081 gggggcggtg gcgaggcgca ggagcttctg gagcactgcg gcgtgtgtcg cgagcgcctg
     1141 cggcccgagc gggatcctcg gctgctgccc tgtctacatt cggcctgcag tgcctgcctg
     1201 ggccccgcta cacccgccgc agcgaataat tcgggggatg gcggctcggc gggcgacggc
     1261 gctagtgagt actgggtggc caggtgcccc tccccctcct cgcagcccgt gctcgggact
     1321 gcgcctgtgc gagagtatgg gggcccgggt agggttaagt aggcctgctg gtttaagagg
     1381 gggcggggaa cgggtcctgg cctctgccaa tgcccgttac caggtctgga caccgaggtg
     1441 cagaatgtga tgggagatgt caagaaatca gagggtgtgc atgcattccc atttttatct
     1501 tggcagtggt ggattgtcca gtgtgcaaac agcagtgcta ctccaaagac atcgtggaga
     1561 attattttat gcgtgatagt ggcagtaagg cctcttctga ttcccaggat gctaaccagg
     1621 tatgtcctgt gggaaatctt ggggaagagg gaagcgtgac cttttccaga gcctctcacc
     1681 atcttcacta gtttggattg tatccttatg cggtctcagt gtctgggagg gtaccgagct
     1741 cgaattcact ggccgtcgtt ttacaacgtc gtgactggga aaaccctggc gttacccaac
     1801 ttaatcgcct tgcagcacat ccccctttcg ccagctggcg taatagcgaa gaggcccgca
     1861 ccgatcgccc ttcccaacag ttgcgcagcc tgaatggcga atggcgcctg atgcggtatt
     1921 ttctccttac gcatctgtgc ggtatttcac accgcatatg gtgcactctc agtacaatct
     1981 gctctgatgc cgcatagtta agccagcccc gacacccgcc aacacccgct gacgcgccct
     2041 gacgggcttg tctgctcccg gcatccgctt acagacaagc tgtgaccgtc tccgggagct
     2101 gcatgtgtca gaggttttca ccgtcatcac cgaaacgcgc gagacgaaag ggcctcgtga
     2161 tacgcctatt tttataggtt aatgtcatga taataatggt ttcttagacg tcaggtggca
     2221 cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata
     2281 tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga
     2341 gtatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca ttttgccttc
     2401 ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat cagttgggtg
     2461 cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag agttttcgcc
     2521 ccgaagaacg ttttccaatg atgagcactt ttaaagttct gctatgtggc gcggtattat
     2581 cccgtattga cgccgggcaa gagcaactcg gtcgccgcat acactattct cagaatgact
     2641 tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca gtaagagaat
     2701 tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt ctgacaacga
     2761 tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat gtaactcgcc
     2821 ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt gacaccacga
     2881 tgcctgtagc aatggcaaca acgttgcgca aactattaac tggcgaacta cttactctag
     2941 cttcccggca acaattaata gactggatgg aggcggataa agttgcagga ccacttctgc
     3001 gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt gagcgtgggt
     3061 ctcgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc gtagttatct
     3121 acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct gagataggtg
     3181 cctcactgat taagcattgg taactgtcag accaagttta ctcatatata ctttagattg
     3241 atttaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttttt gataatctca
     3301 tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc gtagaaaaga
     3361 tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa
     3421 aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga
     3481 aggtaactgg cttcagcaga gcgcagatac caaatactgt tcttctagtg tagccgtagt
     3541 taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt
     3601 taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat
     3661 agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct
     3721 tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca
     3781 cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag
     3841 agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc
     3901 gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga
     3961 aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca
     4021 tgttctttcc tgcgttatcc cctgattctg tggataaccg tattaccgcc tttgagtgag
     4081 ctgataccgc tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc gaggaagcgg
     4141 aagagcgccc aatacgcaaa ccgcctctcc ccgcgcgttg gccgattcat taatgcagct
     4201 ggcacgacag gtttcccgac tggaaagcgg gcagtgagcg caacgcaatt aatgtgagtt
     4261 agctcactca ttaggcaccc caggctttac actttatgct tccggctcgt atgttgtgtg
     4321 gaattgtgag cggataacaa tttcacacag gaaacagcta tgaccatgat tacgccaagc
     4381 ttgcatgcct gcaggtcgac tctagaggat ccccccgagt ttaacaataa gaaagtttgg
     4441 gttaagaccg cctccacgtt cggcgggaac ttcctggcaa agaccccctc accccctcca
     4501 cctgggctcc ggcgctttgc tctgccgtgg ccccgcccac cctgtattgt cccgcccctc
     4561 ttccccgact ataggccagg ctgccgctgc agacgtcgcc acgcctgctg aggaggcggg
     4621 gcctaaggtt cggctctgca ggacctgcgc gctcaagcac gcgggttgcc agcctgggtg
     4681 caccgtgcgg tgggcatggg agacgtcaag cacgcactct gcgggtgaat tcctcaccag
     4741 cagcattgct actctggatc tagaggggcg gcagcgagtg gaagggaggg ccctggcgcg
     4801 tgcgtaatct gcgccccgcc cccaggctgc ggtgcccaac gcgcgtgcgc ggcgtcgtcg
     4861 gcgccagata tttctgtccc gcctccaggc cgcggctctt tctgcgagcg gcgcgcggtc
     4921 aggcggttgt tcgcaagtct tgcggcgcga gtgtctcaga agtgagggag gaggcggcgc
     4981 tggtggcggc gccgtcgtct gtacagaggc ccgcagccag cggcttggcc gcatctcgac
     5041 agcgcctggg gcgcggcggg cgtcggccca ggagacgcgt ggcggcgctc ggcctcgcgg
     5101 catcggcggc tgcctggccg ttggcggcga gcgcacttgc gcctgcgcag cgggctccgt
     5161 gcccctcctc ccctgggcgg cccccccacc cccccggcgg cgtgtgaatg gcggccatgg
     5221 tgagcaaggg cgaggagctg ttcaccgggg tggtgcccat cctggtcgag ctggacggcg
     5281 acgtaaacgg ccacaggttc agcgtgtccg gcgagggcga gggcgatgcc acctacggca
     5341 agctgaccct gaagttcatc tgcaccaccg gcaagctgcc cgtgccctgg cccaccctcg
     5401 tgaccaccct gacctggggc gtgcagtgct tcgcccgcta ccccgaccac atgaagcagc
     5461 acgacttctt caagtccgcc atgcccgaag gctacgtcca ggag
//
