LOCUS       sequence-249272-        6779 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 6779)
  TITLE     Lewin Lab Plasmids
REFERENCE   2  (bases 1 to 6779)
  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-209    C-Tag miRFP670[2] L10 FRB EGFP
FEATURES             Location/Qualifiers
     source          1..6779
                     /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             1034..1309
                     /codon_start=1
                     /product="FKBP-rapamycin binding domain of human FRAP"
                     /label=FRB
                     /translation="ILWHEMWHEGLEEASRLYFGERNVKGMFEVLEPLHAMMERGPQTL
                     KETSFNQAYGRDLMEAQEWCRKYMKSGNVKDLTQAWDLYYHVFRRIS"
     misc_feature    join(1938..2512,2513..2515,2516..2524)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(2105..2122)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     2332..2351
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             join(2525..2527,2528..2530,2531..3244)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(2525..2527,2528..2530,2531..3241)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2570..2591)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2831..2850)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3178..3199
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     intron          3290..3422
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     misc_feature    3431..4019
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3484..3504)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3902..3913)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(4046..4063)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4052..4163
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(4259..4279)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(4259..4277)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(4298..4314)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4298..4314)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4311..4333)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     complement(4416..4435)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    4481..4536
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(4535..4554)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     rep_origin      complement(4915..5503)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4995..5014)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5674..6465,6466..6534))
                     /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     6297..6316
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6535..6639)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6707..6725
                     /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 gccatcctct ggcatgagat gtggcatgaa ggcctggaag aggcatctcg
     1081 tttgtacttt ggggaaagga acgtgaaagg catgtttgag gtgctggagc ccttgcatgc
     1141 tatgatggaa cggggccccc agactctgaa ggaaacatcc tttaatcagg cctatggtcg
     1201 agatttaatg gaggcccaag agtggtgcag gaagtacatg aaatcaggga atgtcaagga
     1261 cctcacccaa gcctgggacc tctattatca tgtgttccga cgaatctcag ggagctctgc
     1321 cagcggtggc agcaccagtg gcagagcggg acccagtatc gacctgtctg ggacactcgc
     1381 accagcactg gagcgcataa gaactgctgg atcactgagg gccctctgtg acgacacagt
     1441 cttgctcttt cagcaatgta cgggctatga tcgagtgatg gtgtaccgat tcgacgaaca
     1501 gggtcatggt ctggtctttt ccgagtgtca cgtgcctggc ctggagagct actttggcaa
     1561 ccgctatccc tcttctaccg taccccagat ggccaggcaa ctttacgtcc ggcaacgggt
     1621 cagagtgctg gtggatgtga cctaccagcc tgtgccactc gaacctcggc tgagcccact
     1681 cactggacgg gatctggaca tgagcggctg cttcctgagg tccatgtcac cttgccatct
     1741 gcagttcttg aaggacatgg gcgttcgagc cacactcgca gtctcactgg ttgtaggcgg
     1801 gaaactgtgg ggtcttgtgg tgtgtcacca ctaccttccc aggttcatcc ggttcgagtt
     1861 gagagccatc tgcaaacgtt tggccgagag aatagccacc cgcattactg ctctggaatc
     1921 ctaagaattc agatccgccc ctctccctcc ccccccccta acgttactgg ccgaagccgc
     1981 ttggaataag gccggtgtgc gtttgtctat atgttatttt ccaccatatt gccgtctttt
     2041 ggcaatgtga gggcccggaa acctggccct gtcttcttga cgagcattcc taggggtctt
     2101 tcccctctcg ccaaaggaat gcaaggtctg ttgaatgtcg tgaaggaagc agttcctctg
     2161 gtagcttctt gaagacaaac aacgtctgta gcgacccttt gcaggcagcg gaacccccca
     2221 cctggcgaca ggtgcctctg cggccaaaag ccacgtgtat aagatacacc tgcaaaggcg
     2281 gcacaacccc agtgccacgt tgtgagttgg atagttgtgg aaagagtcaa atggctctcc
     2341 tcaagcgtat tcaacaaggg gctgaaggat gcccagaagg taccccattg tatgggatct
     2401 gatctggggc ctcggtacac atgctttaca tgtgtttagt cgaggttaaa aaaacgtcta
     2461 ggccccccga accacgggga cgtggttttc ctttgaaaaa cacgatgata atatggccac
     2521 aaccatggtg agcaagggcg aggagctgtt caccggggtg gtgcccatcc tggtcgagct
     2581 ggacggcgac gtaaacggcc acaagttcag cgtgtccggc gagggcgagg gcgatgccac
     2641 ctacggcaag ctgaccctga agttcatctg caccaccggc aagctgcccg tgccctggcc
     2701 caccctcgtg accaccctga cctacggcgt gcagtgcttc agccgctacc ccgaccacat
     2761 gaagcagcac gacttcttca agtccgccat gcccgaaggc tacgtccagg agcgcaccat
     2821 cttcttcaag gacgacggca actacaagac ccgcgccgag gtgaagttcg agggcgacac
     2881 cctggtgaac cgcatcgagc tgaagggcat cgacttcaag gaggacggca acatcctggg
     2941 gcacaagctg gagtacaact acaacagcca caacgtctat atcatggccg acaagcagaa
     3001 gaacggcatc aaggtgaact tcaagatccg ccacaacatc gaggacggca gcgtgcagct
     3061 cgccgaccac taccagcaga acacccccat cggcgacggc cccgtgctgc tgcccgacaa
     3121 ccactacctg agcacccagt ccgccctgag caaagacccc aacgagaagc gcgatcacat
     3181 ggtcctgctg gagttcgtga ccgccgccgg gatcactctc ggcatggacg agctgtacaa
     3241 gtaaaccggt gctcgaggta ccgatgaata gctaaggtcg aggccgcagg taagtatcaa
     3301 ggttacaaga caggtttaag gagaccaata gaaactgggc ttgtcgagac agagaagact
     3361 cttgcgtttc tgataggcac ctattggtct tactgacatc cactttgcct ttctctccac
     3421 aggtgtcgac aatcaacctc tggattacaa aatttgtgaa agattgactg gtattcttaa
     3481 ctatgttgct ccttttacgc tatgtggata cgctgcttta atgcctttgt atcatgctat
     3541 tgcttcccgt atggctttca ttttctcctc cttgtataaa tcctggttgc tgtctcttta
     3601 tgaggagttg tggcccgttg tcaggcaacg tggcgtggtg tgcactgtgt ttgctgacgc
     3661 aacccccact ggttggggca ttgccaccac ctgtcagctc ctttccggga ctttcgcttt
     3721 ccccctccct attgccacgg cggaactcat cgccgcctgc cttgcccgct gctggacagg
     3781 ggctcggctg ttgggcactg acaattccgt ggtgttgtcg gggaagctga cgtcctttcc
     3841 atggctgctc gcctgtgttg ccacctggat tctgcgcggg acgtccttct gctacgtccc
     3901 ttcggccctc aatccagcgg accttccttc ccgcggcctg ctgccggctc tgcggcctct
     3961 tccgcgtctt cgccttcgcc ctcagacgag tcggatctcc ctttgggccg cctccccgcc
     4021 tggaattcga gctcggtacg atcagcctcg actgtgcctt ctagttgcca gccatctgtt
     4081 gtttgcccct cccccgtgcc ttccttgacc ctggaaggtg ccactcccac tgtcctttcc
     4141 taataaaatg aggaaattgc atcgcattgt ctgagtaggt gtcattctat tctggggggt
     4201 ggggtggggc aggacagcaa gggggaggat tgggaagaca atagcccagc ttttgttccc
     4261 tttagtgagg gttaattgcg cgcttggcgt aatcatggtc atagctgttt cctgtgtgaa
     4321 attgttatcc gctaattcac tcctcaggtg caggctgcct atcagaaggt ggtggctggt
     4381 gtggccaatg ccctggctca caaataccac tgagatcttt ttccctctgc caaaaattat
     4441 ggggacatca tgaagcccct tgagcatctg acttctggct aataaaggaa atttattttc
     4501 attgcaatag tgtgttggaa ttttttgtgt ctctcactcg gaaggacata tgggagggca
     4561 aatcatttaa aacatcagaa tgagtatttg gtttagagtt tggcaacata tgcccatatg
     4621 ctggctgcca tgaacaaagg ttggctataa agaggtcatc agtatatgaa acagccccct
     4681 gctgtccatt ccttattcca tagaaaagcc ttgacttgag gttagatttt ttttatattt
     4741 tgttttgtgt tatttttttc tttaacatcc ctaaaatttt ccttacatgt tttactagcc
     4801 agatttttcc tcctctcctg actactccca gtcatagctg tccctcttct cttatggaga
     4861 tcggccggcc gccggcgtcg acgcgcgcgg ccgcggcgcg ccgttgctgg cgtttttcca
     4921 taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa
     4981 cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc
     5041 tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc
     5101 gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct
     5161 gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg
     5221 tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag
     5281 gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta
     5341 cggctacact agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg
     5401 aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt
     5461 tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt
     5521 ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag
     5581 attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat
     5641 ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc
     5701 tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat
     5761 aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc
     5821 acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag
     5881 aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag
     5941 agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt
     6001 ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg
     6061 agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt
     6121 tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc
     6181 tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc
     6241 attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa
     6301 taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg
     6361 aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc
     6421 caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag
     6481 gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt
     6541 cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt
     6601 tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc
     6661 acctgacgtc taagaaacca ttattatcat gacattaacc tataaaaata ggcgtatcac
     6721 gaggcccttt cgtctacgcg cgcgccggcg gcgcgccgtc gacggccggc cgcggccgc
//
