LOCUS       sequence-364583-        5498 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For knocking in FKBP F36V P2A mRuby2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5498)
  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 5498)
  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.mRuby2
FEATURES             Location/Qualifiers
     source          1..5498
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             84..794
                     /codon_start=1
                     /product="monomeric red fluorescent protein derived from 
                     mRuby, with improved photophysical properties (Lam et al., 
                     2012)"
                     /label=mRuby2
                     /translation="MVSKGEELIKENMRMKVVMEGSVNGHQFKCTGEGEGNPYMGTQTM
                     RIKVIEGGPLPFAFDILATSFMYGSRTFIKYPKGIPDFFKQSFPEGFTWERVTRYEDGG
                     VVTVMQDTSLEDGCLVYHVQVRGVNFPSNGPVMQKKTKGWEPNTEMMYPADGGLRGYTH
                     MALKVDGGGHLSCSFVTTYRSKKTVGNIKMPGIHAVDHRLERLEESDNEMFVVQREHAV
                     AKFAGLGGGMDELYK"
     CDS             804..860
                     /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             861..887
                     /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             861..887
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             891..917
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             927..1247
                     /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(2113..2130)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2113..2129)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2122..2144)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(2338..2357)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2457..2479
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2517..2535)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2603..2707
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2708..2776,2777..3568)
                     /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(2926..2945)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3739..4327
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4228..4247
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4481..4498
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    4615..4636
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(4651..4656,4657..4674,4675..4681)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4689..4705
                     /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     4694..4716
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     4713..4729
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4713..4729
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
ORIGIN
        1 cacttgcgcc tgcgcagcgg gctccgtgcc cctcctcccc tgggcggccc ccccaccccc
       61 ccggcggcgt gtgaatggcg gccatggtgt ctaagggcga agagctgatc aaggaaaata
      121 tgcgtatgaa ggtggtcatg gaaggttcgg tcaacggcca ccaattcaaa tgcacaggtg
      181 aaggagaagg caatccgtac atgggaactc aaaccatgag gatcaaagtc atcgagggag
      241 gacccctgcc atttgccttt gacattcttg ccacgtcgtt catgtatggc agccgtactt
      301 ttatcaagta cccgaaaggc attcctgatt tctttaaaca gtcctttcct gagggtttta
      361 cttgggaaag agttacgaga tacgaagatg gtggagtcgt caccgtcatg caggacacca
      421 gccttgagga tggctgtctc gtttaccacg tccaagtcag aggggtaaac tttccctcca
      481 atggtcccgt gatgcagaag aagaccaagg gttgggagcc taatacagag atgatgtatc
      541 cagcagatgg tggtctgagg ggatacactc atatggcact gaaagttgat ggtggtggcc
      601 atctgtcttg ctctttcgta acaacttaca ggtcaaaaaa gaccgtcggg aacatcaaga
      661 tgcccggtat ccatgccgtt gatcaccgcc tggaaaggtt agaggaaagt gacaatgaaa
      721 tgttcgtagt acaacgcgaa cacgcagttg ccaagttcgc cgggcttggt ggtgggatgg
      781 acgagctgta caagggaagc ggagctacta acttcagcct gctgaagcag gctggagacg
      841 tggaggagaa ccctggacct tacccctacg acgtgcccga ctacgccggc tatccgtatg
      901 atgtcccgga ctatgcagga agcggaggag tgcaggtgga aaccatctcc ccaggagacg
      961 ggcgcacctt ccccaagcgc ggccagacct gcgtggtgca ctacaccggg atgcttgaag
     1021 atggaaagaa agttgattcc tcccgggaca gaaacaagcc ctttaagttt atgctaggca
     1081 agcaggaggt gatccgaggc tgggaagaag gggttgccca gatgagtgtg ggtcagagag
     1141 ccaaactgac tatatctcca gattatgcct atggtgccac tgggcaccca ggcatcatcc
     1201 caccacatgc cactctcgtc ttcgatgtgg agcttctaaa actggaaggt ggcggtggct
     1261 cgggcggtgg tgggtcgggt ggcggcggat ctgcctcggc ggcagcgact gcagcggcct
     1321 cggccgcgac ggccgcctcg gcggcctctg gtagcccagg gtcgggcgag ggctcggcgg
     1381 gcggtgagaa gcgtccggct gcttcctcag ccgcggcggc ctctgcagcc gcgtcgtccc
     1441 ctgcgggggg cggtggcgag gcgcaggagc ttctggagca ctgcggcgtg tgtcgcgagc
     1501 gcctgcggcc cgagcgggat cctcggctgc tgccctgtct acattcggcc tgcagtgcct
     1561 gcctgggccc cgctacaccc gccgcagcga ataattcggg ggatggcggc tcggcgggcg
     1621 acggcgctag tgagtactgg gtggccaggt gcccctcccc ctcctcgcag cccgtgctcg
     1681 ggactgcgcc tgtgcgagag tatgggggcc cgggtagggt taagtaggcc tgctggttta
     1741 agagggggcg gggaacgggt cctggcctct gccaatgccc gttaccaggt ctggacaccg
     1801 aggtgcagaa tgtgatggga gatgtcaaga aatcagaggg tgtgcatgca ttcccatttt
     1861 tatcttggca gtggtggatt gtccagtgtg caaacagcag tgctactcca aagacatcgt
     1921 ggagaattat tttatgcgtg atagtggcag taaggcctct tctgattccc aggatgctaa
     1981 ccaggtatgt cctgtgggaa atcttgggga agagggaagc gtgacctttt ccagagcctc
     2041 tcaccatctt cactagtttg gattgtatcc ttatgcggtc tcagtgtctg ggagggtacc
     2101 gagctcgaat tcactggccg tcgttttaca acgtcgtgac tgggaaaacc ctggcgttac
     2161 ccaacttaat cgccttgcag cacatccccc tttcgccagc tggcgtaata gcgaagaggc
     2221 ccgcaccgat cgcccttccc aacagttgcg cagcctgaat ggcgaatggc gcctgatgcg
     2281 gtattttctc cttacgcatc tgtgcggtat ttcacaccgc atatggtgca ctctcagtac
     2341 aatctgctct gatgccgcat agttaagcca gccccgacac ccgccaacac ccgctgacgc
     2401 gccctgacgg gcttgtctgc tcccggcatc cgcttacaga caagctgtga ccgtctccgg
     2461 gagctgcatg tgtcagaggt tttcaccgtc atcaccgaaa cgcgcgagac gaaagggcct
     2521 cgtgatacgc ctatttttat aggttaatgt catgataata atggtttctt agacgtcagg
     2581 tggcactttt cggggaaatg tgcgcggaac ccctatttgt ttatttttct aaatacattc
     2641 aaatatgtat ccgctcatga gacaataacc ctgataaatg cttcaataat attgaaaaag
     2701 gaagagtatg agtattcaac atttccgtgt cgcccttatt cccttttttg cggcattttg
     2761 ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta aaagatgctg aagatcagtt
     2821 gggtgcacga gtgggttaca tcgaactgga tctcaacagc ggtaagatcc ttgagagttt
     2881 tcgccccgaa gaacgttttc caatgatgag cacttttaaa gttctgctat gtggcgcggt
     2941 attatcccgt attgacgccg ggcaagagca actcggtcgc cgcatacact attctcagaa
     3001 tgacttggtt gagtactcac cagtcacaga aaagcatctt acggatggca tgacagtaag
     3061 agaattatgc agtgctgcca taaccatgag tgataacact gcggccaact tacttctgac
     3121 aacgatcgga ggaccgaagg agctaaccgc ttttttgcac aacatggggg atcatgtaac
     3181 tcgccttgat cgttgggaac cggagctgaa tgaagccata ccaaacgacg agcgtgacac
     3241 cacgatgcct gtagcaatgg caacaacgtt gcgcaaacta ttaactggcg aactacttac
     3301 tctagcttcc cggcaacaat taatagactg gatggaggcg gataaagttg caggaccact
     3361 tctgcgctcg gcccttccgg ctggctggtt tattgctgat aaatctggag ccggtgagcg
     3421 tgggtctcgc ggtatcattg cagcactggg gccagatggt aagccctccc gtatcgtagt
     3481 tatctacacg acggggagtc aggcaactat ggatgaacga aatagacaga tcgctgagat
     3541 aggtgcctca ctgattaagc attggtaact gtcagaccaa gtttactcat atatacttta
     3601 gattgattta aaacttcatt tttaatttaa aaggatctag gtgaagatcc tttttgataa
     3661 tctcatgacc aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag accccgtaga
     3721 aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac
     3781 aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt
     3841 tccgaaggta actggcttca gcagagcgca gataccaaat actgttcttc tagtgtagcc
     3901 gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat
     3961 cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag
     4021 acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc
     4081 cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag
     4141 cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac
     4201 aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg
     4261 gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct
     4321 atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc
     4381 tcacatgttc tttcctgcgt tatcccctga ttctgtggat aaccgtatta ccgcctttga
     4441 gtgagctgat accgctcgcc gcagccgaac gaccgagcgc agcgagtcag tgagcgagga
     4501 agcggaagag cgcccaatac gcaaaccgcc tctccccgcg cgttggccga ttcattaatg
     4561 cagctggcac gacaggtttc ccgactggaa agcgggcagt gagcgcaacg caattaatgt
     4621 gagttagctc actcattagg caccccaggc tttacacttt atgcttccgg ctcgtatgtt
     4681 gtgtggaatt gtgagcggat aacaatttca cacaggaaac agctatgacc atgattaccc
     4741 caagcttgca tgcctgcagg tcgactctag aggatccccc cgagtttaac aataagaaag
     4801 tttgggttaa gaccgcctcc acgttcggcg ggaacttcct ggcaaagacc ccctcacccc
     4861 ctccacctgg gctccggcgc tttgctctgc cgtggccccg cccaccctgt attgtcccgc
     4921 ccctcttccc cgactatagg ccaggctgcc gctgcagacg tcgccacgcc tgctgaggag
     4981 gcggggccta aggttcggct ctgcaggacc tgcgcgctca agcacgcggg ttgccagcct
     5041 gggtgcaccg tgcggtgggc atgggagacg tcaagcacgc actctgcggg tgaattcctc
     5101 accagcagca ttgctactct ggatctagag gggcggcagc gagtggaagg gagggccctg
     5161 gcgcgtgcgt aatctgcgcc ccgcccccag gctgcggtgc ccaacgcgcg tgcgcggcgt
     5221 cgtcggcgcc agatatttct gtcccgcctc caggccgcgg ctctttctgc gagcggcgcg
     5281 cggtcaggcg gttgttcgca agtcttgcgg cgcgagtgtc tcagaagcga gggaggaggc
     5341 ggcgctggtg gcggcgccgt cgtctgtaca gaggcccgca gccagcggct tggccgcatc
     5401 tcgacagcgc ctggggcgcg gcgggcgtcg gcccaggaga cgcgtggcgg cgctcggcct
     5461 cgcggcatcg gcggctgcct ggccgttggc ggcgagcg
//
