LOCUS       sequence-197859-        5650 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For knocking in FKBP F36V P2A BFP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5650)
  AUTHORS   Weintraub AS, Li CH, Zamudio AV, Sigova AA, Hannett NM, Day DS, 
            Abraham BJ, Cohen MA, Nabet B, Buckley DL, Guo YE, Hnisz D, Jaenisch
            R, Bradner JE, Gray NS, Young RA
  TITLE     YY1 Is a Structural Regulator of Enhancer-Promoter Loops.
  JOURNAL   Cell. 2017 Dec 14;171(7):1573-1588.e28. doi: 
            10.1016/j.cell.2017.11.008. Epub 2017 Dec 7.
  PUBMED    29224777
REFERENCE   2  (bases 1 to 5650)
  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: 
            "10.1016/j.cell.2017.11.008"; journalName: "Cell"; date: 
            "2017-12-14- 14"; volume: "171"; issue: "7"; pages: "1573-1588.e28"
COMMENT     Sequence Label: pAW63.YY1.FKBP.knock-in.BFP
FEATURES             Location/Qualifiers
     source          1..5650
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    106..127
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(142..147,148..165,166..172)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    180..196
                     /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     185..207
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     204..220
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     204..220
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             1166..1486
                     /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"
     CDS             1493..1519
                     /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             1493..1519
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             1523..1549
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             1559..1615
                     /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             1616..2323
                     /codon_start=1
                     /product="enhanced monomeric blue fluorescent protein 
                     (Subach et al., 2011)"
                     /label=mTagBFP2
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELIKENMHMKLYMEGTVDNHHFKCTSEGEGKPYEGTQTMR
                     IKVVEGGPLPFAFDILATSFLYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGV
                     LTATQDTSLQDGCLIYNVKIRGVNFTSNGPVMQKKTLGWEAFTETLYPADGGLEGRNDM
                     ALKLVGGSHLIANAKTTYRSKKPAKNLKMPGVYYVDYRLERIKEANNETYVEQHEVAVA
                     RYCDLPSKLGHKLN"
     primer_bind     complement(3254..3271)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3254..3270)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3263..3285)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(3479..3498)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3598..3620
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3658..3676)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3744..3848
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3849..3917,3918..4709)
                     /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(4067..4086)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4880..5468
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5369..5388
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5622..5639
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca
       61 cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaatg tgagttagct
      121 cactcattag gcaccccagg ctttacactt tatgcttccg gctcgtatgt tgtgtggaat
      181 tgtgagcgga taacaatttc acacaggaaa cagctatgac catgattacg ccaagcttgc
      241 atgcctgcag gtcgacaagc cctttcaggt agagccagta cctgttcccc acactgaaag
      301 ctagggtgct gggcagggtg gtgatatcaa gtaccatggg gcgcctgtgg gggcatctta
      361 ttcccactcc cacttccata gctctctgaa ggtctgcttg ggttcctggt tacttactgc
      421 atgtgactgc aggtgttaca tactagggga tgggagtatc tgtcgggaga aacagggtaa
      481 tgtgagtgag agaaccccgc actaaattcc tagcatgttt acagagactc agagattgta
      541 tgtttggtag tgagtagtgt gttgaattac tgagagagtg ctcataagca ctgttggcta
      601 catctccagg ggacacttgg tgctgcttat agaaaaatca agtgagtctg tgacagttgg
      661 catcactaaa tatcttaaat catctgtatt tattctactt tctggagtgt ctctgaaaat
      721 actcaggcct taaatttgca cttagtacaa cccttctgtt tttgtaagag ctaaccatgt
      781 gttcatagtg tcagagatcc ctgtgtgtca gttctaaatc acgctcagaa gggtctccag
      841 aggaggaagg agacacactc actctagtac agtctagtct agtacagtgg ttttccttcc
      901 aagcagtgag ccattgaccg cccctgattc tggtcccgtt tgccaagtgg gttgatgtct
      961 ggtttttcct tgacagtgca cattcgaagg ctgcgggaag cgcttttcac tggacttcaa
     1021 tttgcgcaca catgtgcgaa tccataccgg agacaggccc tatgtgtgcc ccttcgacgg
     1081 ttgtaataag aagtttgctc agtcaactaa cctgaaatct cacatcttaa cacacgctaa
     1141 agccaaaaac aaccagggat ccggaggagt gcaggtggaa accatctccc caggagacgg
     1201 gcgcaccttc cccaagcgcg gccagacctg cgtggtgcac tacaccggga tgcttgaaga
     1261 tggaaagaaa gttgattcct cccgggacag aaacaagccc tttaagttta tgctaggcaa
     1321 gcaggaggtg atccgaggct gggaagaagg ggttgcccag atgagtgtgg gtcagagagc
     1381 caaactgact atatctccag attatgccta tggtgccact gggcacccag gcatcatccc
     1441 accacatgcc actctcgtct tcgatgtgga gcttctaaaa ctggaaggcg gctaccccta
     1501 cgacgtgccc gactacgccg gctatccgta tgatgtcccg gactatgcag gctccggagc
     1561 aacaaacttc tctctgctga aacaagccgg agatgtcgaa gagaatcctg gaccggtgtc
     1621 taagggcgaa gagctgatta aggagaacat gcacatgaag ctgtacatgg agggcaccgt
     1681 ggacaaccat cacttcaagt gcacatccga gggcgaaggc aagccctacg agggcaccca
     1741 gaccatgaga atcaaggtgg tcgagggcgg ccctctcccc ttcgccttcg acatcctggc
     1801 tactagcttc ctctacggca gcaagacctt catcaaccac acccagggca tccccgactt
     1861 cttcaagcag tccttccctg agggcttcac atgggagaga gtcaccacat acgaagacgg
     1921 gggcgtgctg accgctaccc aggacaccag cctccaggac ggctgcctca tctacaacgt
     1981 caagatcaga ggggtgaact tcacatccaa cggccctgtg atgcagaaga aaacactcgg
     2041 ctgggaggcc ttcaccgaga cgctgtaccc cgctgacggc ggcctggaag gcagaaacga
     2101 catggccctg aagctcgtgg gcgggagcca tctgatcgca aacgccaaga ccacatatag
     2161 atccaagaaa cccgctaaga acctcaagat gcctggcgtc tactatgtgg actacagact
     2221 ggaaagaatc aaggaggcca acaacgagac ctacgtcgag cagcacgagg tggcagtggc
     2281 cagatactgc gacctcccta gcaaactggg gcacaagctt aattaaaaag aagagagaag
     2341 accttctcga cccgggaagc ctcttcagga gtgtgattgg gaataaatat gcctctcctt
     2401 tgtatattat ttctaggaag aattttaaaa atgaatccta cacacttaag ggacatgttt
     2461 tgataaagta gtaaaaattt aaaaaatact ttaataagat gacattgcta agatgctata
     2521 tcttgctctg taatctcgtt tcaaaaacaa ggtgtttttg taaagtgtgg tcccaacagg
     2581 aggacaattc atgaacttcg catcaaaaga caattcttta tacaacagtg ctaaaaatgg
     2641 gacttctttt cacattctta taaatatgaa gctcacctgt tgcttacaat ttttttaatt
     2701 ttgtattttc caagtgtgca tattgtacac tttttgggga tatgcttagt aatgctgtgt
     2761 gattttctgg aggttgataa ctttgcttgc ggtagatttt ctttaaaaga atgggcagtt
     2821 acatgcatac ttcaaaagta tttttcctgt acaaaaaaaa agttatatag gttttgtttg
     2881 ctatcttaat tttggttgta ttctttgatg ttaacacatt ttgtataatt gtatcgtata
     2941 gctgtattga atcatgtaga atcaaatatt agatgtgatt taatagtgtt aatcaattta
     3001 aacccatttt agtcactttt ttttccccaa aaaatactgc cagatgctga tgttcagtgt
     3061 aatttctttg cctgttcagt tacagaaagt ggtgctcagt tgtagaatgt attgtacctt
     3121 ttaacatctg atgtgtacat ccgtgtaaca gaaagggcaa caataaaata gcgatcctaa
     3181 agaaagatta cggcagaaag agctctgtaa gcacagcctt attttcggat ccccgggtac
     3241 cgagctcgaa ttcactggcc gtcgttttac aacgtcgtga ctgggaaaac cctggcgtta
     3301 cccaacttaa tcgccttgca gcacatcccc ctttcgccag ctggcgtaat agcgaagagg
     3361 cccgcaccga tcgcccttcc caacagttgc gcagcctgaa tggcgaatgg cgcctgatgc
     3421 ggtattttct ccttacgcat ctgtgcggta tttcacaccg catatggtgc actctcagta
     3481 caatctgctc tgatgccgca tagttaagcc agccccgaca cccgccaaca cccgctgacg
     3541 cgccctgacg ggcttgtctg ctcccggcat ccgcttacag acaagctgtg accgtctccg
     3601 ggagctgcat gtgtcagagg ttttcaccgt catcaccgaa acgcgcgaga cgaaagggcc
     3661 tcgtgatacg cctattttta taggttaatg tcatgataat aatggtttct tagacgtcag
     3721 gtggcacttt tcggggaaat gtgcgcggaa cccctatttg tttatttttc taaatacatt
     3781 caaatatgta tccgctcatg agacaataac cctgataaat gcttcaataa tattgaaaaa
     3841 ggaagagtat gagtattcaa catttccgtg tcgcccttat tccctttttt gcggcatttt
     3901 gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt
     3961 tgggtgcacg agtgggttac atcgaactgg atctcaacag cggtaagatc cttgagagtt
     4021 ttcgccccga agaacgtttt ccaatgatga gcacttttaa agttctgcta tgtggcgcgg
     4081 tattatcccg tattgacgcc gggcaagagc aactcggtcg ccgcatacac tattctcaga
     4141 atgacttggt tgagtactca ccagtcacag aaaagcatct tacggatggc atgacagtaa
     4201 gagaattatg cagtgctgcc ataaccatga gtgataacac tgcggccaac ttacttctga
     4261 caacgatcgg aggaccgaag gagctaaccg cttttttgca caacatgggg gatcatgtaa
     4321 ctcgccttga tcgttgggaa ccggagctga atgaagccat accaaacgac gagcgtgaca
     4381 ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact attaactggc gaactactta
     4441 ctctagcttc ccggcaacaa ttaatagact ggatggaggc ggataaagtt gcaggaccac
     4501 ttctgcgctc ggcccttccg gctggctggt ttattgctga taaatctgga gccggtgagc
     4561 gtgggtctcg cggtatcatt gcagcactgg ggccagatgg taagccctcc cgtatcgtag
     4621 ttatctacac gacggggagt caggcaacta tggatgaacg aaatagacag atcgctgaga
     4681 taggtgcctc actgattaag cattggtaac tgtcagacca agtttactca tatatacttt
     4741 agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc ctttttgata
     4801 atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag
     4861 aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa
     4921 caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt
     4981 ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgttctt ctagtgtagc
     5041 cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa
     5101 tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa
     5161 gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc
     5221 ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa
     5281 gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa
     5341 caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg
     5401 ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc
     5461 tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg
     5521 ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg
     5581 agtgagctga taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg
     5641 aagcggaaga
//
