LOCUS       sequence-249292-        6476 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  PPI (BiFC/TriFC) assay vector.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6476)
  TITLE     Lewin Lab Plasmids
REFERENCE   2  (bases 1 to 6476)
  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"
COMMENT     Sequence Label: pRDAX-471    C-Tag UnaG[1] L20 FRB EBPF2
FEATURES             Location/Qualifiers
     source          1..6476
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        152..531
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        532..735
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     686..706
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     732..756
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     intron          871..1003
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     regulatory      1018..1027
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      1022..1031
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1033..1308
                     /codon_start=1
                     /product="FKBP-rapamycin binding domain of human FRAP"
                     /label=FRB
                     /translation="ILWHEMWHEGLEEASRLYFGERNVKGMFEVLEPLHAMMERGPQTL
                     KETSFNQAYGRDLMEAQEWCRKYMKSGNVKDLTQAWDLYYHVFRRIS"
     misc_feature    join(1634..2208,2209..2211,2212..2220)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(1801..1818)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     2028..2047
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             join(2221..2223,2224..2226,2227..2940)
                     /codon_start=1
                     /product="enhanced blue variant of GFP (Ai et al., 2007)"
                     /label=EBFP2
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTL
                     KFICTTGKLPVPWPTLVTTLSHGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GTYKTRAEVKFEGDTLVNRIELKGVDFKEDGNILGHKLEYNFNSHNIYIMAVKQKNGIK
                     VNFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDSHYLSTQSVLSKDPNEKRDHMVLL
                     EFRTAAGITLGMDELYK"
     primer_bind     complement(2266..2287)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     intron          2986..3118
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     misc_feature    3127..3715
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3180..3200)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3598..3609)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(3742..3759)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3748..3859
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(3955..3975)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(3955..3973)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3994..4010)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3994..4010)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4007..4029)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     complement(4112..4131)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    4177..4232
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(4231..4250)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     rep_origin      complement(4611..5199)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4691..4710)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5370..6161,6162..6230))
                     /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     5993..6012
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6231..6335)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6403..6421
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 cctgcctcca ggcctcaata ttggccatta gccatattat tcattggtta tatagcataa
       61 atcaatattg gctattggcc attgcatacg ttgtatctat atcataatat gtacatttat
      121 attggctcat gtccaatatg accgccatgt tggcattgat tattgactag ttattaatag
      181 taatcaatta cggggtcatt agttcatagc ccatatatgg agttccgcgt tacataactt
      241 acggtaaatg gcccgcctgg ctgaccgccc aacgaccccc gcccattgac gtcaataatg
      301 acgtatgttc ccatagtaac gccaataggg actttccatt gacgtcaatg ggtggagtat
      361 ttacggtaaa ctgcccactt ggcagtacat caagtgtatc atatgccaag tccgccccct
      421 attgacgtca atgacggtaa atggcccgcc tggcattatg cccagtacat gaccttacgg
      481 gactttccta cttggcagta catctacgta ttagtcatcg ctattaccat ggtgatgcgg
      541 ttttggcagt acaccaatgg gcgtggatag cggtttgact cacggggatt tccaagtctc
      601 caccccattg acgtcaatgg gagtttgttt tggcaccaaa atcaacggga ctttccaaaa
      661 tgtcgtaata accccgcccc gttgacgcaa atgggcggta ggcgtgtacg gtgggaggtc
      721 tatataagca gagctcgttt agtgaaccgt cagatcacta gaagctttat tgcggtagtt
      781 tatcacagtt aaattgctaa cgcagtcagt gcttctgaca caacagtctc gaacttaagc
      841 tgcagaagtt ggtcgtgagg cactgggcag gtaagtatca aggttacaag acaggtttaa
      901 ggagaccaat agaaactggg cttgtcgaga cagagaagac tcttgcgttt ctgataggca
      961 cctattggtc ttactgacat ccactttgcc tttctctcca caggtgtcca ctcccaggcc
     1021 accatggtgg ccatcctctg gcatgagatg tggcatgaag gcctggaaga ggcatctcgt
     1081 ttgtactttg gggaaaggaa cgtgaaaggc atgtttgagg tgctggagcc cttgcatgct
     1141 atgatggaac ggggccccca gactctgaag gaaacatcct ttaatcaggc ctatggtcga
     1201 gatttaatgg aggcccaaga gtggtgcagg aagtacatga aatcagggaa tgtcaaggac
     1261 ctcacccaag cctgggacct ctattatcat gtgttccgac gaatctcagg gagtcctggc
     1321 ggtggatccg gaggctctgg gagctctgcc agcggtggca gcaccagtgt cgaaaagttc
     1381 gtgggcacat ggaagattgc cgatagccac aacttcggcg aatacctcaa agctataggg
     1441 gcaccaaaag agcttagcga tggtggagat gcgacaactc ccacactgta cattagccag
     1501 aaagacggag acaagatgac tgtcaagatc gagaatgggc ctcccacctt tctggacaca
     1561 caggttaagt tcaaactggg tgaggagttt gacgagtttc ccagtgatcg caggaaataa
     1621 gaattcagat ccgcccctct ccctcccccc cccctaacgt tactggccga agccgcttgg
     1681 aataaggccg gtgtgcgttt gtctatatgt tattttccac catattgccg tcttttggca
     1741 atgtgagggc ccggaaacct ggccctgtct tcttgacgag cattcctagg ggtctttccc
     1801 ctctcgccaa aggaatgcaa ggtctgttga atgtcgtgaa ggaagcagtt cctctggtag
     1861 cttcttgaag acaaacaacg tctgtagcga ccctttgcag gcagcggaac cccccacctg
     1921 gcgacaggtg cctctgcggc caaaagccac gtgtataaga tacacctgca aaggcggcac
     1981 aaccccagtg ccacgttgtg agttggatag ttgtggaaag agtcaaatgg ctctcctcaa
     2041 gcgtattcaa caaggggctg aaggatgccc agaaggtacc ccattgtatg ggatctgatc
     2101 tggggcctcg gtacacatgc tttacatgtg tttagtcgag gttaaaaaaa cgtctaggcc
     2161 ccccgaacca cggggacgtg gttttccttt gaaaaacacg atgataatat ggccacaacc
     2221 atggtgagca agggcgagga gctgttcacc ggggtggtgc ccatcctggt cgagctggac
     2281 ggcgacgtaa acggccacaa gttcagcgtg aggggcgagg gcgagggcga tgccaccaac
     2341 ggcaagctga ccctgaagtt catctgcacc accggcaagc tgcccgtgcc ctggcccacc
     2401 ctcgtgacca ccctgagcca cggcgtgcag tgcttcgccc gctaccccga ccacatgaag
     2461 cagcacgact tcttcaagtc cgccatgccc gaaggctacg tccaggagcg caccatcttc
     2521 ttcaaggacg acggcaccta caagacccgc gccgaggtga agttcgaggg cgacaccctg
     2581 gtgaaccgca tcgagctgaa gggcgtcgac ttcaaggagg acggcaacat cctggggcac
     2641 aagctggagt acaacttcaa cagccacaac atctatatca tggccgtcaa gcagaagaac
     2701 ggcatcaagg tgaacttcaa gatccgccac aacgtggagg acggcagcgt gcagctcgcc
     2761 gaccactacc agcagaacac ccccatcggc gacggccccg tgctgctgcc cgacagccac
     2821 tacctgagca cccagtccgt gctgagcaaa gaccccaacg agaagcgcga tcacatggtc
     2881 ctgctggagt tccgcaccgc cgccgggatc actctcggca tggacgagct gtacaagtaa
     2941 accggtgctc gaggtaccga tgaatagcta aggtcgaggc cgcaggtaag tatcaaggtt
     3001 acaagacagg tttaaggaga ccaatagaaa ctgggcttgt cgagacagag aagactcttg
     3061 cgtttctgat aggcacctat tggtcttact gacatccact ttgcctttct ctccacaggt
     3121 gtcgacaatc aacctctgga ttacaaaatt tgtgaaagat tgactggtat tcttaactat
     3181 gttgctcctt ttacgctatg tggatacgct gctttaatgc ctttgtatca tgctattgct
     3241 tcccgtatgg ctttcatttt ctcctccttg tataaatcct ggttgctgtc tctttatgag
     3301 gagttgtggc ccgttgtcag gcaacgtggc gtggtgtgca ctgtgtttgc tgacgcaacc
     3361 cccactggtt ggggcattgc caccacctgt cagctccttt ccgggacttt cgctttcccc
     3421 ctccctattg ccacggcgga actcatcgcc gcctgccttg cccgctgctg gacaggggct
     3481 cggctgttgg gcactgacaa ttccgtggtg ttgtcgggga agctgacgtc ctttccatgg
     3541 ctgctcgcct gtgttgccac ctggattctg cgcgggacgt ccttctgcta cgtcccttcg
     3601 gccctcaatc cagcggacct tccttcccgc ggcctgctgc cggctctgcg gcctcttccg
     3661 cgtcttcgcc ttcgccctca gacgagtcgg atctcccttt gggccgcctc cccgcctgga
     3721 attcgagctc ggtacgatca gcctcgactg tgccttctag ttgccagcca tctgttgttt
     3781 gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc ctttcctaat
     3841 aaaatgagga aattgcatcg cattgtctga gtaggtgtca ttctattctg gggggtgggg
     3901 tggggcagga cagcaagggg gaggattggg aagacaatag cccagctttt gttcccttta
     3961 gtgagggtta attgcgcgct tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg
     4021 ttatccgcta attcactcct caggtgcagg ctgcctatca gaaggtggtg gctggtgtgg
     4081 ccaatgccct ggctcacaaa taccactgag atctttttcc ctctgccaaa aattatgggg
     4141 acatcatgaa gccccttgag catctgactt ctggctaata aaggaaattt attttcattg
     4201 caatagtgtg ttggaatttt ttgtgtctct cactcggaag gacatatggg agggcaaatc
     4261 atttaaaaca tcagaatgag tatttggttt agagtttggc aacatatgcc catatgctgg
     4321 ctgccatgaa caaaggttgg ctataaagag gtcatcagta tatgaaacag ccccctgctg
     4381 tccattcctt attccataga aaagccttga cttgaggtta gatttttttt atattttgtt
     4441 ttgtgttatt tttttcttta acatccctaa aattttcctt acatgtttta ctagccagat
     4501 ttttcctcct ctcctgacta ctcccagtca tagctgtccc tcttctctta tggagatcgg
     4561 ccggccgccg gcgtcgacgc gcgcggccgc ggcgcgccgt tgctggcgtt tttccatagg
     4621 ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg
     4681 acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt
     4741 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt
     4801 tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc
     4861 tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt
     4921 gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt
     4981 agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc
     5041 tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa
     5101 agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt
     5161 tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct
     5221 acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta
     5281 tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa
     5341 agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc
     5401 tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact
     5461 acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc
     5521 tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt
     5581 ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta
     5641 agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg
     5701 tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt
     5761 acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc
     5821 agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt
     5881 actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc
     5941 tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc
     6001 gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa
     6061 ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac
     6121 tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa
     6181 aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt
     6241 tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa
     6301 tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct
     6361 gacgtctaag aaaccattat tatcatgaca ttaacctata aaaataggcg tatcacgagg
     6421 ccctttcgtc tacgcgcgcg ccggcggcgc gccgtcgacg gccggccgcg gccgcg
//
