LOCUS       sequence-372428-        5420 bp    DNA     circular SYN 14-APR-2025
DEFINITION  Expresses His-tagged, Arabinose-inducible mCherry-XL (R125K) in 
            bacteria.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5420)
  AUTHORS   Mukherjee S, Manna P, Hung ST, Vietmeyer F, Friis P, Palmer AE, 
            Jimenez R
  TITLE     Directed Evolution of a Bright Variant of mCherry: Suppression of 
            Nonradiative Decay by Fluorescence Lifetime Selections.
  JOURNAL   J Phys Chem B. 2022 Jun 16. doi: 10.1021/acs.jpcb.2c01956.
  PUBMED    35709514
REFERENCE   2  (bases 1 to 5420)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "J Phys Chem
            B. 2022 Jun 16. doi: 10.1021/acs.jpcb.2c01956."
COMMENT     Sequence Label: pBad-mCherry-XL (R125K)
FEATURES             Location/Qualifiers
     source          1..5420
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..285
                     /gene="araBAD"
                     /label=araBAD promoter
                     /note="promoter of the L-arabinose operon of E. coli; the 
                     araC regulatory gene is transcribed in the opposite 
                     direction (Guzman et al., 1995)"
     primer_bind     208..227
                     /label=pBAD Forward
                     /note="For vectors with E. coli araBAD promoter, forward 
                     primer"
     RBS             312..334
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             354..371
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             375..407
                     /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(411..419,420..434)
                     /codon_start=1
                     /product="Xpress(TM) epitope tag, including an enterokinase
                     recognition and cleavage site"
                     /label=Xpress(TM) tag
                     /translation="DLYDDDDK"
     CDS             447..1157
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLKGTNFPSDGPVMQKKTMGSEASSERMYPEDGALK
                     GEVKYRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNRKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(592..610)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     860..879
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     complement(1203..1220)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      1373..1459
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      1551..1578
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        1597..1688
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1689..1757,1758..2549)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(1907..1926)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2591..3046
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2678..2697)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2888..2909
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     rep_origin      3157..3745
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3646..3665
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3899..3916
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3931..4071
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3997..4016)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4157..4179
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     4317..4336
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     CDS             complement(4516..5394)
                     /codon_start=1
                     /gene="araC"
                     /product="L-arabinose regulatory protein"
                     /label=araC
                     /translation="MAEAQNDPLLPGYSFNAHLVAGLTPIEANGYLDFFIDRPLGMKGY
                     ILNLTIRGQGVVKNQGREFVCRPGDILLFPPGEIHHYGRHPEAREWYHQWVYFRPRAYW
                     HEWLNWPSIFANTGFFRPDEAHQPHFSDLFGQIINAGQGEGRYSELLAINLLEQLLLRR
                     MEAINESLHPPMDNRVREACQYISDHLADSNFDIASVAQHVCLSPSRLSHLFRQQLGIS
                     VLSWREDQRISQAKLLLSTTRMPIATVGRNVGFDDQLYFSRVFKKCTGASPSEFRAGCE
                     EKVNDVAVKLS"
ORIGIN
        1 aagaaaccaa ttgtccatat tgcatcagac attgccgtca ctgcgtcttt tactggctct
       61 tctcgctaac caaaccggta accccgctta ttaaaagcat tctgtaacaa agcgggacca
      121 aagccatgac aaaaacgcgt aacaaaagtg tctataatca cggcagaaaa gtccacattg
      181 attatttgca cggcgtcaca ctttgctatg ccatagcatt tttatccata agattagcgg
      241 atactacctg acgcttttta tcgcaactct ctactgtttc tccatacccg tttttttggg
      301 ctagaaataa ttttgtttaa ctttaagaag gagatataca tatgcggggt tctcatcatc
      361 atcatcatca tggtatggct agcatgactg gtggacagca aatgggtcgg gatctgtacg
      421 acgatgacga taaggatccc aaaaaaatgg tgagcaaggg cgaggaggat aacatggcca
      481 tcatcaagga gttcatgcgc ttcaaggtgc acatggaggg ctccgtgaac ggccacgagt
      541 tcgagatcga gggcgagggc gagggccgcc cctacgaggg cacccagacc gccaagctga
      601 aggtgaccaa gggtggcccc ctgcccttcg cctgggacat cctgtcccct cagttcatgt
      661 acggctccaa ggcctacgtg aagcaccccg ccgacatccc cgactacttg aagctgtcat
      721 tccccgaggg cttcaagtgg gagcgcgtga tgaacttcga ggacggcggc gtggtgaccg
      781 tgacccagga ctcctccctg caggacggcg agttcatcta caaggtgaag ctgaagggca
      841 ccaactttcc ctccgacggc cccgtaatgc agaagaagac catgggcagt gaggcctcct
      901 ccgagcggat gtaccccgag gacggcgccc tgaagggcga ggttaagtat aggctgaagc
      961 tgaaggacgg cggccactac gacgctgagg tcaagaccac ctacaaggcc aagaagcccg
     1021 tgcagctgcc cggcgcctac aacgtcaacc gtaagttgga catcacctcc cacaacgagg
     1081 actacaccat cgtggaacag tacgaacgcg ccgagggccg ccactccacc ggcggcatgg
     1141 acgagctgta caagtaagaa ttcgaagctt ggctgttttg gcggatgaga gaagattttc
     1201 agcctgatac agattaaatc agaacgcaga agcggtctga taaaacagaa tttgtctggc
     1261 ggcagtagcg cggtggtccc acctgacccc atgccgaact cagaagtgaa acgccgtagc
     1321 gccgatggta gtgtggggtc tccccatgcg agagtaggga actgccaggc atcaaataaa
     1381 acgaaaggct cagtcgaaag actgggcctt tcgttttatc tgttgtttgt cggtgaacgc
     1441 tctccggagt aggacaaatc cgccgggagc ggatttgaac gttgcgaagc aacggcccgg
     1501 agggtggcgg gcaggacgcc cgccataaac tgccaggcat caaattaagc agaaggccat
     1561 cctgacggat ggcctttttg cgtttctaca aactcttttg tttatttttc taaatacatt
     1621 caaatatgta tccgctcatg agacaataac cctgataaat gcttcaataa tattgaaaaa
     1681 ggaagagtat gagtattcaa catttccgtg tcgcccttat tccctttttt gcggcatttt
     1741 gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt
     1801 tgggtgcacg agtgggttac atcgaactgg atctcaacag cggtaagatc cttgagagtt
     1861 ttcgccccga agaacgtttt ccaatgatga gcacttttaa agttctgcta tgtggcgcgg
     1921 tattatcccg tgttgacgcc gggcaagagc aactcggtcg ccgcatacac tattctcaga
     1981 atgacttggt tgagtactca ccagtcacag aaaagcatct tacggatggc atgacagtaa
     2041 gagaattatg cagtgctgcc ataaccatga gtgataacac tgcggccaac ttacttctga
     2101 caacgatcgg aggaccgaag gagctaaccg cttttttgca caacatgggg gatcatgtaa
     2161 ctcgccttga tcgttgggaa ccggagctga atgaagccat accaaacgac gagcgtgaca
     2221 ccacgatgcc tgcagcaatg gcaacaacgt tgcgcaaact attaactggc gaactactta
     2281 ctctagcttc ccggcaacaa ttaatagact ggatggaggc ggataaagtt gcaggaccac
     2341 ttctgcgctc ggcccttccg gctggctggt ttattgctga taaatctgga gccggtgagc
     2401 gtgggtctcg cggtatcatt gcagcactgg ggccagatgg taagccctcc cgtatcgtag
     2461 ttatctacac gacggggagt caggcaacta tggatgaacg aaatagacag atcgctgaga
     2521 taggtgcctc actgattaag cattggtaac tgtcagacca agtttactca tatatacttt
     2581 agattgattt acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc
     2641 agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc
     2701 tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg
     2761 ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgatttggg tgatggttca
     2821 cgtagtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc
     2881 tttaatagtg gactcttgtt ccaaactgga acaacactca accctatctc gggctattct
     2941 tttgatttat aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa
     3001 caaaaattta acgcgaattt taacaaaata ttaacgttta caatttaaaa ggatctaggt
     3061 gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg
     3121 agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt
     3181 aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca
     3241 agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac
     3301 tgtccttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcatac
     3361 atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct
     3421 taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg
     3481 gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga gatacctaca
     3541 gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt
     3601 aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta
     3661 tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc
     3721 gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc
     3781 cttttgctgg ccttttgctc acatgttctt tcctgcgtta tcccctgatt ctgtggataa
     3841 ccgtattacc gcctttgagt gagctgatac cgctcgccgc agccgaacga ccgagcgcag
     3901 cgagtcagtg agcgaggaag cggaagagcg cctgatgcgg tattttctcc ttacgcatct
     3961 gtgcggtatt tcacaccgca tctggtgcac tctcagtaca atctgctctg atgccgcata
     4021 gttaagccag tatacactcc gctatcgcta cgtgactggg tcatggctgc gccccgacac
     4081 ccgccaacac ccgctgacgc gccctgacgg gcttgtctgc tcccggcatc cgcttacaga
     4141 caagctgtga ccgtctccgg gagctgcatg tgtcagaggt tttcaccgtc atcaccgaaa
     4201 cgcgcgaggc agaaggagat ggcgcccaac agtcccccgg ccacggggcc tgccaccata
     4261 cccacgccga aacaagcgct catgagcccg aagtggcgag cccgatcttc cccatcggtg
     4321 atgtcggcga tataggcgcc agcaaccgca cctgtggcgc cggtgatgcc ggccacgatg
     4381 cgtccggcgt agaggatctg ctcatgtttg acagcttatc atcgatgcat aatgtgcctg
     4441 tcaaatggac gaagcaggga ttctgcaaac cctatgctac tccgtcaagc cgtcaattgt
     4501 ctgattcgtt accaattatg acaacttgac ggctacatca ttcacttttt cttcacaacc
     4561 ggcacggaac tcgctcgggc tggccccggt gcatttttta aatacccgcg agaaatagag
     4621 ttgatcgtca aaaccaacat tgcgaccgac ggtggcgata ggcatccggg tggtgctcaa
     4681 aagcagcttc gcctggctga tacgttggtc ctcgcgccag cttaagacgc taatccctaa
     4741 ctgctggcgg aaaagatgtg acagacgcga cggcgacaag caaacatgct gtgcgacgct
     4801 ggcgatatca aaattgctgt ctgccaggtg atcgctgatg tactgacaag cctcgcgtac
     4861 ccgattatcc atcggtggat ggagcgactc gttaatcgct tccatgcgcc gcagtaacaa
     4921 ttgctcaagc agatttatcg ccagcagctc cgaatagcgc ccttcccctt gcccggcgtt
     4981 aatgatttgc ccaaacaggt cgctgaaatg cggctggtgc gcttcatccg ggcgaaagaa
     5041 ccccgtattg gcaaatattg acggccagtt aagccattca tgccagtagg cgcgcggacg
     5101 aaagtaaacc cactggtgat accattcgcg agcctccgga tgacgaccgt agtgatgaat
     5161 ctctcctggc gggaacagca aaatatcacc cggtcggcaa acaaattctc gtccctgatt
     5221 tttcaccacc ccctgaccgc gaatggtgag attgagaata taacctttca ttcccagcgg
     5281 tcggtcgata aaaaaatcga gataaccgtt ggcctcaatc ggcgttaaac ccgccaccag
     5341 atgggcatta aacgagtatc ccggcagcag gggatcattt tgcgcttcag ccatactttt
     5401 catactcccg ccattcagag
//
