LOCUS       sequence-249217-        6588 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 6588)
  TITLE     Lewin Lab Plasmids
REFERENCE   2  (bases 1 to 6588)
  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-336    N-Tag miRFP670[2] L32 {Esp3I(2x)}...
FEATURES             Location/Qualifiers
     source          1..6588
                     /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"
     misc_feature    join(1747..2321,2322..2324,2325..2333)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(1914..1931)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     2141..2160
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             join(2334..2336,2337..2339,2340..3053)
                     /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(2334..2336,2337..2339,2340..3050)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2379..2400)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2640..2659)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2987..3008
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     intron          3099..3231
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     misc_feature    3240..3828
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3293..3313)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3711..3722)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(3855..3872)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3861..3972
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(4068..4088)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(4068..4086)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(4107..4123)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4107..4123)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4120..4142)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     complement(4225..4244)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    4290..4345
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(4344..4363)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     rep_origin      complement(4724..5312)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4804..4823)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5483..6274,6275..6343))
                     /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     6106..6125
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6344..6448)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6516..6534
                     /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 caccatgggc agagcgggac ccagtatcga cctgtctggg acactcgcac cagcactgga
     1081 gcgcataaga actgctggat cactgagggc cctctgtgac gacacagtct tgctctttca
     1141 gcaatgtacg ggctatgatc gagtgatggt gtaccgattc gacgaacagg gtcatggtct
     1201 ggtcttttcc gagtgtcacg tgcctggcct ggagagctac tttggcaacc gctatccctc
     1261 ttctaccgta ccccagatgg ccaggcaact ttacgtccgg caacgggtca gagtgctggt
     1321 ggatgtgacc taccagcctg tgccactcga acctcggctg agcccactca ctggacggga
     1381 tctggacatg agcggctgct tcctgaggtc catgtcacct tgccatctgc agttcttgaa
     1441 ggacatgggc gttcgagcca cactcgcagt ctcactggtt gtaggcggga aactgtgggg
     1501 tcttgtggtg tgtcaccact accttcccag gttcatccgg ttcgagttga gagccatctg
     1561 caaacgtttg gccgagagaa tagccacccg cattactgct ctggaatccg acgtgggtgg
     1621 cggagggagc gaaggcggcg gatccggagg gcctggctct ggcggagagg ggagcgctgg
     1681 aggcgggtca gccggtggtg ggagtcgaga cgccgaggca cgccgtctcc taagaattca
     1741 gatccgcccc tctccctccc ccccccctaa cgttactggc cgaagccgct tggaataagg
     1801 ccggtgtgcg tttgtctata tgttattttc caccatattg ccgtcttttg gcaatgtgag
     1861 ggcccggaaa cctggccctg tcttcttgac gagcattcct aggggtcttt cccctctcgc
     1921 caaaggaatg caaggtctgt tgaatgtcgt gaaggaagca gttcctctgg tagcttcttg
     1981 aagacaaaca acgtctgtag cgaccctttg caggcagcgg aaccccccac ctggcgacag
     2041 gtgcctctgc ggccaaaagc cacgtgtata agatacacct gcaaaggcgg cacaacccca
     2101 gtgccacgtt gtgagttgga tagttgtgga aagagtcaaa tggctctcct caagcgtatt
     2161 caacaagggg ctgaaggatg cccagaaggt accccattgt atgggatctg atctggggcc
     2221 tcggtacaca tgctttacat gtgtttagtc gaggttaaaa aaacgtctag gccccccgaa
     2281 ccacggggac gtggttttcc tttgaaaaac acgatgataa tatggccaca accatggtga
     2341 gcaagggcga ggagctgttc accggggtgg tgcccatcct ggtcgagctg gacggcgacg
     2401 taaacggcca caagttcagc gtgtccggcg agggcgaggg cgatgccacc tacggcaagc
     2461 tgaccctgaa gttcatctgc accaccggca agctgcccgt gccctggccc accctcgtga
     2521 ccaccctgac ctacggcgtg cagtgcttca gccgctaccc cgaccacatg aagcagcacg
     2581 acttcttcaa gtccgccatg cccgaaggct acgtccagga gcgcaccatc ttcttcaagg
     2641 acgacggcaa ctacaagacc cgcgccgagg tgaagttcga gggcgacacc ctggtgaacc
     2701 gcatcgagct gaagggcatc gacttcaagg aggacggcaa catcctgggg cacaagctgg
     2761 agtacaacta caacagccac aacgtctata tcatggccga caagcagaag aacggcatca
     2821 aggtgaactt caagatccgc cacaacatcg aggacggcag cgtgcagctc gccgaccact
     2881 accagcagaa cacccccatc ggcgacggcc ccgtgctgct gcccgacaac cactacctga
     2941 gcacccagtc cgccctgagc aaagacccca acgagaagcg cgatcacatg gtcctgctgg
     3001 agttcgtgac cgccgccggg atcactctcg gcatggacga gctgtacaag taaaccggtg
     3061 ctcgaggtac cgatgaatag ctaaggtcga ggccgcaggt aagtatcaag gttacaagac
     3121 aggtttaagg agaccaatag aaactgggct tgtcgagaca gagaagactc ttgcgtttct
     3181 gataggcacc tattggtctt actgacatcc actttgcctt tctctccaca ggtgtcgaca
     3241 atcaacctct ggattacaaa atttgtgaaa gattgactgg tattcttaac tatgttgctc
     3301 cttttacgct atgtggatac gctgctttaa tgcctttgta tcatgctatt gcttcccgta
     3361 tggctttcat tttctcctcc ttgtataaat cctggttgct gtctctttat gaggagttgt
     3421 ggcccgttgt caggcaacgt ggcgtggtgt gcactgtgtt tgctgacgca acccccactg
     3481 gttggggcat tgccaccacc tgtcagctcc tttccgggac tttcgctttc cccctcccta
     3541 ttgccacggc ggaactcatc gccgcctgcc ttgcccgctg ctggacaggg gctcggctgt
     3601 tgggcactga caattccgtg gtgttgtcgg ggaagctgac gtcctttcca tggctgctcg
     3661 cctgtgttgc cacctggatt ctgcgcggga cgtccttctg ctacgtccct tcggccctca
     3721 atccagcgga ccttccttcc cgcggcctgc tgccggctct gcggcctctt ccgcgtcttc
     3781 gccttcgccc tcagacgagt cggatctccc tttgggccgc ctccccgcct ggaattcgag
     3841 ctcggtacga tcagcctcga ctgtgccttc tagttgccag ccatctgttg tttgcccctc
     3901 ccccgtgcct tccttgaccc tggaaggtgc cactcccact gtcctttcct aataaaatga
     3961 ggaaattgca tcgcattgtc tgagtaggtg tcattctatt ctggggggtg gggtggggca
     4021 ggacagcaag ggggaggatt gggaagacaa tagcccagct tttgttccct ttagtgaggg
     4081 ttaattgcgc gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg
     4141 ctaattcact cctcaggtgc aggctgccta tcagaaggtg gtggctggtg tggccaatgc
     4201 cctggctcac aaataccact gagatctttt tccctctgcc aaaaattatg gggacatcat
     4261 gaagcccctt gagcatctga cttctggcta ataaaggaaa tttattttca ttgcaatagt
     4321 gtgttggaat tttttgtgtc tctcactcgg aaggacatat gggagggcaa atcatttaaa
     4381 acatcagaat gagtatttgg tttagagttt ggcaacatat gcccatatgc tggctgccat
     4441 gaacaaaggt tggctataaa gaggtcatca gtatatgaaa cagccccctg ctgtccattc
     4501 cttattccat agaaaagcct tgacttgagg ttagattttt tttatatttt gttttgtgtt
     4561 atttttttct ttaacatccc taaaattttc cttacatgtt ttactagcca gatttttcct
     4621 cctctcctga ctactcccag tcatagctgt ccctcttctc ttatggagat cggccggccg
     4681 ccggcgtcga cgcgcgcggc cgcggcgcgc cgttgctggc gtttttccat aggctccgcc
     4741 cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac
     4801 tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc
     4861 tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata
     4921 gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc
     4981 acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca
     5041 acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag
     5101 cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta
     5161 gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg
     5221 gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc
     5281 agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt
     5341 ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa
     5401 ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat
     5461 atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga
     5521 tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac
     5581 gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg
     5641 ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg
     5701 caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt
     5761 cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct
     5821 cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat
     5881 cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta
     5941 agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca
     6001 tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat
     6061 agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac
     6121 atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa
     6181 ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt
     6241 cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg
     6301 caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat
     6361 attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt
     6421 agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtct
     6481 aagaaaccat tattatcatg acattaacct ataaaaatag gcgtatcacg aggccctttc
     6541 gtctacgcgc gcgccggcgg cgcgccgtcg acggccggcc gcggccgc
//
