LOCUS       sequence-249269-        6602 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 6602)
  TITLE     Lewin Lab Plasmids
REFERENCE   2  (bases 1 to 6602)
  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-206    N-Tag miRFP670[1] L20 FRB EBPF2
FEATURES             Location/Qualifiers
     source          1..6602
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        153..532
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        533..736
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     687..707
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     733..757
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     intron          872..1004
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     regulatory      1019..1028
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      1023..1032
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1469..1744
                     /codon_start=1
                     /product="FKBP-rapamycin binding domain of human FRAP"
                     /label=FRB
                     /translation="ILWHEMWHEGLEEASRLYFGERNVKGMFEVLEPLHAMMERGPQTL
                     KETSFNQAYGRDLMEAQEWCRKYMKSGNVKDLTQAWDLYYHVFRRIS"
     misc_feature    join(1761..2335,2336..2338,2339..2347)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(1928..1945)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     2155..2174
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             join(2348..2350,2351..2353,2354..3067)
                     /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(2393..2414)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     intron          3113..3245
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     misc_feature    3254..3842
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3307..3327)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3725..3736)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(3869..3886)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3875..3986
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(4082..4102)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(4082..4100)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(4121..4137)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4121..4137)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4134..4156)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     complement(4239..4258)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    4304..4359
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(4358..4377)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     rep_origin      complement(4738..5326)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4818..4837)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5497..6288,6289..6357))
                     /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     6120..6139
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6358..6462)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6530..6548
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 gcctgcctcc aggcctcaat attggccatt agccatatta ttcattggtt atatagcata
       61 aatcaatatt ggctattggc cattgcatac gttgtatcta tatcataata tgtacattta
      121 tattggctca tgtccaatat gaccgccatg ttggcattga ttattgacta gttattaata
      181 gtaatcaatt acggggtcat tagttcatag cccatatatg gagttccgcg ttacataact
      241 tacggtaaat ggcccgcctg gctgaccgcc caacgacccc cgcccattga cgtcaataat
      301 gacgtatgtt cccatagtaa cgccaatagg gactttccat tgacgtcaat gggtggagta
      361 tttacggtaa actgcccact tggcagtaca tcaagtgtat catatgccaa gtccgccccc
      421 tattgacgtc aatgacggta aatggcccgc ctggcattat gcccagtaca tgaccttacg
      481 ggactttcct acttggcagt acatctacgt attagtcatc gctattacca tggtgatgcg
      541 gttttggcag tacaccaatg ggcgtggata gcggtttgac tcacggggat ttccaagtct
      601 ccaccccatt gacgtcaatg ggagtttgtt ttggcaccaa aatcaacggg actttccaaa
      661 atgtcgtaat aaccccgccc cgttgacgca aatgggcggt aggcgtgtac ggtgggaggt
      721 ctatataagc agagctcgtt tagtgaaccg tcagatcact agaagcttta ttgcggtagt
      781 ttatcacagt taaattgcta acgcagtcag tgcttctgac acaacagtct cgaacttaag
      841 ctgcagaagt tggtcgtgag gcactgggca ggtaagtatc aaggttacaa gacaggttta
      901 aggagaccaa tagaaactgg gcttgtcgag acagagaaga ctcttgcgtt tctgataggc
      961 acctattggt cttactgaca tccactttgc ctttctctcc acaggtgtcc actcccaggc
     1021 caccatggtg gctgggcatg ctagcggttc tcctgccttt ggcaccgcta gtcactccaa
     1081 ctgcgagcac gaggagatcc atcttgctgg gagtatccag cctcacggag cccttctggt
     1141 cgttagcgaa cacgatcacc gtgtgattca ggcaagcgcg aatgcagccg agttcctcaa
     1201 cctgggcagt gtgctgggag ttcccttggc cgaaatcgac ggagatctgc tgatcaagat
     1261 tctgccccat ctcgatccga cagctgaagg catgccagta gcagttaggt gtcgcattgg
     1321 gaatccgtcc accgagtatt gcggtctgat gcataggcca ccagaaggag ggctgattat
     1381 cgagctggaa agagcgggac ccgggagtcc tggcggtgga tccggaggct ctgggagctc
     1441 tgccagcggt ggcagcacct ccgtggccat cctctggcat gagatgtggc atgaaggcct
     1501 ggaagaggca tctcgtttgt actttgggga aaggaacgtg aaaggcatgt ttgaggtgct
     1561 ggagcccttg catgctatga tggaacgggg cccccagact ctgaaggaaa catcctttaa
     1621 tcaggcctat ggtcgagatt taatggaggc ccaagagtgg tgcaggaagt acatgaaatc
     1681 agggaatgtc aaggacctca cccaagcctg ggacctctat tatcatgtgt tccgacgaat
     1741 ctcataagaa ttcagatccg cccctctccc tccccccccc ctaacgttac tggccgaagc
     1801 cgcttggaat aaggccggtg tgcgtttgtc tatatgttat tttccaccat attgccgtct
     1861 tttggcaatg tgagggcccg gaaacctggc cctgtcttct tgacgagcat tcctaggggt
     1921 ctttcccctc tcgccaaagg aatgcaaggt ctgttgaatg tcgtgaagga agcagttcct
     1981 ctggtagctt cttgaagaca aacaacgtct gtagcgaccc tttgcaggca gcggaacccc
     2041 ccacctggcg acaggtgcct ctgcggccaa aagccacgtg tataagatac acctgcaaag
     2101 gcggcacaac cccagtgcca cgttgtgagt tggatagttg tggaaagagt caaatggctc
     2161 tcctcaagcg tattcaacaa ggggctgaag gatgcccaga aggtacccca ttgtatggga
     2221 tctgatctgg ggcctcggta cacatgcttt acatgtgttt agtcgaggtt aaaaaaacgt
     2281 ctaggccccc cgaaccacgg ggacgtggtt ttcctttgaa aaacacgatg ataatatggc
     2341 cacaaccatg gtgagcaagg gcgaggagct gttcaccggg gtggtgccca tcctggtcga
     2401 gctggacggc gacgtaaacg gccacaagtt cagcgtgagg ggcgagggcg agggcgatgc
     2461 caccaacggc aagctgaccc tgaagttcat ctgcaccacc ggcaagctgc ccgtgccctg
     2521 gcccaccctc gtgaccaccc tgagccacgg cgtgcagtgc ttcgcccgct accccgacca
     2581 catgaagcag cacgacttct tcaagtccgc catgcccgaa ggctacgtcc aggagcgcac
     2641 catcttcttc aaggacgacg gcacctacaa gacccgcgcc gaggtgaagt tcgagggcga
     2701 caccctggtg aaccgcatcg agctgaaggg cgtcgacttc aaggaggacg gcaacatcct
     2761 ggggcacaag ctggagtaca acttcaacag ccacaacatc tatatcatgg ccgtcaagca
     2821 gaagaacggc atcaaggtga acttcaagat ccgccacaac gtggaggacg gcagcgtgca
     2881 gctcgccgac cactaccagc agaacacccc catcggcgac ggccccgtgc tgctgcccga
     2941 cagccactac ctgagcaccc agtccgtgct gagcaaagac cccaacgaga agcgcgatca
     3001 catggtcctg ctggagttcc gcaccgccgc cgggatcact ctcggcatgg acgagctgta
     3061 caagtaaacc ggtgctcgag gtaccgatga atagctaagg tagaggccgc aggtaagtat
     3121 caaggttaca agacaggttt aaggagacca atagaaactg ggcttgtcga gacagagaag
     3181 actcttgcgt ttctgatagg cacctattgg tcttactgac atccactttg cctttctctc
     3241 cacaggtgtc gacaatcaac ctctggatta caaaatttgt gaaagattga ctggtattct
     3301 taactatgtt gctcctttta cgctatgtgg atacgctgct ttaatgcctt tgtatcatgc
     3361 tattgcttcc cgtatggctt tcattttctc ctccttgtat aaatcctggt tgctgtctct
     3421 ttatgaggag ttgtggcccg ttgtcaggca acgtggcgtg gtgtgcactg tgtttgctga
     3481 cgcaaccccc actggttggg gcattgccac cacctgtcag ctcctttccg ggactttcgc
     3541 tttccccctc cctattgcca cggcggaact catcgccgcc tgccttgccc gctgctggac
     3601 aggggctcgg ctgttgggca ctgacaattc cgtggtgttg tcggggaagc tgacgtcctt
     3661 tccatggctg ctcgcctgtg ttgccacctg gattctgcgc gggacgtcct tctgctacgt
     3721 cccttcggcc ctcaatccag cggaccttcc ttcccgcggc ctgctgccgg ctctgcggcc
     3781 tcttccgcgt cttcgccttc gccctcagac gagtcggatc tccctttggg ccgcctcccc
     3841 gcctggaatt cgagctcggt acgatcagcc tcgactgtgc cttctagttg ccagccatct
     3901 gttgtttgcc cctcccccgt gccttccttg accctggaag gtgccactcc cactgtcctt
     3961 tcctaataaa atgaggaaat tgcatcgcat tgtctgagta ggtgtcattc tattctgggg
     4021 ggtggggtgg ggcaggacag caagggggag gattgggaag acaatagccc agcttttgtt
     4081 ccctttagtg agggttaatt gcgcgcttgg cgtaatcatg gtcatagctg tttcctgtgt
     4141 gaaattgtta tccgctaatt cactcctcag gtgcaggctg cctatcagaa ggtggtggct
     4201 ggtgtggcca atgccctggc tcacaaatac cactgagatc tttttccctc tgccaaaaat
     4261 tatggggaca tcatgaagcc ccttgagcat ctgacttctg gctaataaag gaaatttatt
     4321 ttcattgcaa tagtgtgttg gaattttttg tgtctctcac tcggaaggac atatgggagg
     4381 gcaaatcatt taaaacatca gaatgagtat ttggtttaga gtttggcaac atatgcccat
     4441 atgctggctg ccatgaacaa aggttggcta taaagaggtc atcagtatat gaaacagccc
     4501 cctgctgtcc attccttatt ccatagaaaa gccttgactt gaggttagat tttttttata
     4561 ttttgttttg tgttattttt ttctttaaca tccctaaaat tttccttaca tgttttacta
     4621 gccagatttt tcctcctctc ctgactactc ccagtcatag ctgtccctct tctcttatgg
     4681 agatcggccg gccgccggcg tcgacgcgcg cggccgcggc gcgccgttgc tggcgttttt
     4741 ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg
     4801 aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc
     4861 tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt
     4921 ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa
     4981 gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta
     5041 tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa
     5101 caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa
     5161 ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt
     5221 cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt
     5281 ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat
     5341 cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat
     5401 gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc
     5461 aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc
     5521 acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta
     5581 gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga
     5641 cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg
     5701 cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc
     5761 tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat
     5821 cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag
     5881 gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat
     5941 cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa
     6001 ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa
     6061 gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga
     6121 taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg
     6181 gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc
     6241 acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg
     6301 aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact
     6361 cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat
     6421 atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt
     6481 gccacctgac gtctaagaaa ccattattat catgacatta acctataaaa ataggcgtat
     6541 cacgaggccc tttcgtctac gcgcgcgccg gcggcgcgcc gtcgacggcc ggccgcggcc
     6601 gc
//
