LOCUS       sequence-216915-        3547 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For bacterial expression and purification of mRFP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3547)
  AUTHORS   DeGroot ACM, Busch DJ, Hayden CC, Mihelic SA, Alpar AT, Behar M, 
            Stachowiak JC
  TITLE     Entropic Control of Receptor Recycling Using Engineered Ligands.
  JOURNAL   Biophys J. 2018 Mar 27;114(6):1377-1388. doi: 
            10.1016/j.bpj.2018.01.036.
  PUBMED    29590595
REFERENCE   2  (bases 1 to 3547)
  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.bpj.2018.01"; journalName: "Biophys J"; date: 
            "2018-03-27- 27"; volume: "114"; issue: "6"; pages: "1377-1388"
COMMENT     Sequence Label: pRSET his-mRFP
FEATURES             Location/Qualifiers
     source          1..3547
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     45..64
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        45..63
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             95..117
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             137..154
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             158..190
                     /codon_start=1
                     /product="leader peptide from bacteriophage T7 gene 10"
                     /label=T7 tag (gene 10 leader)
                     /note="promotes efficient translation in E. coli"
                     /translation="MASMTGGQQMG"
     CDS             join(194..202,203..217)
                     /codon_start=1
                     /product="Xpress(TM) epitope tag, including an enterokinase
                     recognition and cleavage site"
                     /label=Xpress(TM) tag
                     /translation="DLYDDDDK"
     CDS             233..910
                     /codon_start=1
                     /product="monomeric derivative of DsRed (Campbell et al., 
                     2002)"
                     /label=mRFP1
                     /translation="MASSEDVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFEDGGV
                     VTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASTERMYPEDGALKGEIKM
                     RLKLKDGGHYDAEVKTTYMAKKPVQLPGAYKTDIKLDITSHNEDYTIVEQYERAEGRHS
                     TGA"
     CDS             233..910
                     /codon_start=1
                     /product="monomeric derivative of DsRed (Campbell et al., 
                     2002)"
                     /label=mRFP1
                     /note="mammalian codon-optimized"
                     /translation="MASSEDVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFEDGGV
                     VTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASTERMYPEDGALKGEIKM
                     RLKLKDGGHYDAEVKTTYMAKKPVQLPGAYKTDIKLDITSHNEDYTIVEQYERAEGRHS
                     TGA"
     primer_bind     complement(363..381)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     631..650
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     825..848
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     primer_bind     complement(973..991)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      987..1034
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1131..1586
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1218..1237)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1428..1449
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        1612..1716
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1717..1785,1786..2577)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPKERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(1935..1954)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2748..3336
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3237..3256
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3490..3507
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 cgcgttggcc gattcattaa tgcaggatct cgatcccgcg aaattaatac gactcactat
       61 agggagacca caacggtttc cctctagaaa taattttgtt taactttaag aaggagatat
      121 acatatgcgg ggttctcatc atcatcatca tcatggtatg gctagcatga ctggtggaca
      181 gcaaatgggt cgggatctgt acgacgatga cgataaggat cgatggggat ccatggcctc
      241 ctccgaggac gtcatcaagg agttcatgcg cttcaaggtg cgcatggagg gctccgtgaa
      301 cggccacgag ttcgagatcg agggcgaggg cgagggccgc ccctacgagg gcacccagac
      361 cgccaagctg aaggtgacca agggcggccc cctgcccttc gcctgggaca tcctgtcccc
      421 tcagttccag tacggctcca aggcctacgt gaagcacccc gccgacatcc ccgactactt
      481 gaagctgtcc ttccccgagg gcttcaagtg ggagcgcgtg atgaacttcg aggacggcgg
      541 cgtggtgacc gtgacccagg actcctccct gcaggacggc gagttcatct acaaggtgaa
      601 gctgcgcggc accaacttcc cctccgacgg ccccgtaatg cagaagaaga ccatgggctg
      661 ggaggcctcc accgagcgga tgtaccccga ggacggcgcc ctgaagggcg agatcaagat
      721 gaggctgaag ctgaaggacg gcggccacta cgacgccgag gtcaagacca cctacatggc
      781 caagaagccc gtgcagctgc ccggcgccta caagaccgac atcaagctgg acatcacctc
      841 ccacaacgag gactacacca tcgtggaaca gtacgagcgc gccgagggcc gccactccac
      901 cggcgcctaa gaattcgaag cttgatccgg ctgctaacaa agcccgaaag gaagctgagt
      961 tggctgctgc caccgctgag caataactag cataacccct tggggcctct aaacgggtct
     1021 tgaggggttt tttgctgaaa ggaggaacta tatccggatc tggcgtaata gcgaagaggc
     1081 ccgcaccgat cgcccttccc aacagttgcg cagcctgaat ggcgaatggg acgcgccctg
     1141 tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc
     1201 cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg
     1261 ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta gtgctttacg
     1321 gcacctcgac cccaaaaaac ttgattaggg tgatggttca cgtagtgggc catcgccctg
     1381 atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg gactcttgtt
     1441 ccaaactgga acaacactca accctatctc ggtctattct tttgatttat aagggatttt
     1501 gccgatttcg gcctattggt taaaaaatga gctgatttaa caaaaattta acgcgaattt
     1561 taacaaaata ttaacgctta caatttaggt ggcacttttc ggggaaatgt gcgcggaacc
     1621 cctatttgtt tatttttcta aatacattca aatatgtatc cgctcatgag acaataaccc
     1681 tgataaatgc ttcaataata ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc
     1741 gcccttattc ccttttttgc ggcattttgc cttcctgttt ttgctcaccc agaaacgctg
     1801 gtgaaagtaa aagatgctga agatcagttg ggtgcacgag tgggttacat cgaactggat
     1861 ctcaacagcg gtaagatcct tgagagtttt cgccccaaag aacgttttcc aatgatgagc
     1921 acttttaaag ttctgctatg tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa
     1981 ctcggtcgcc gcatacacta ttctcagaat gacttggttg agtactcacc agtcacagaa
     2041 aagcatctta cggatggcat gacagtaaga gaattatgca gtgctgccat aaccatgagt
     2101 gataacactg cggccaactt acttctgaca acgatcggag gaccgaagga gctaaccgct
     2161 tttttgcaca acatggggga tcatgtaact cgccttgatc gttgggaacc ggagctgaat
     2221 gaagccatac caaacgacga gcgtgacacc acgatgcctg tagcaatggc aacaacgttg
     2281 cgcaaactat taactggcga actacttact ctagcttccc ggcaacaatt aatagactgg
     2341 atggaggcgg ataaagttgc aggaccactt ctgcgctcgg cccttccggc tggctggttt
     2401 attgctgata aatctggagc cggtgagcgt gggtctcgcg gtatcattgc agcactgggg
     2461 ccagatggta agccctcccg tatcgtagtt atctacacga cggggagtca ggcaactatg
     2521 gatgaacgaa atagacagat cgctgagata ggtgcctcac tgattaagca ttggtaactg
     2581 tcagaccaag tttactcata tatactttag attgatttaa aacttcattt ttaatttaaa
     2641 aggatctagg tgaagatcct ttttgataat ctcatgacca aaatccctta acgtgagttt
     2701 tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt
     2761 tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt
     2821 ttgccggatc aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag
     2881 ataccaaata ctgttcttct agtgtagccg tagttaggcc accacttcaa gaactctgta
     2941 gcaccgccta catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat
     3001 aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg
     3061 ggctgaacgg ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg
     3121 agatacctac agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac
     3181 aggtatccgg taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga
     3241 aacgcctggt atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt
     3301 ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta
     3361 cggttcctgg ccttttgctg gccttttgct cacatgttct ttcctgcgtt atcccctgat
     3421 tctgtggata accgtattac cgcctttgag tgagctgata ccgctcgccg cagccgaacg
     3481 accgagcgca gcgagtcagt gagcgaggaa gcggaagagc gcccaatacg caaaccgcct
     3541 ctccccg
//
