LOCUS       sequence-476070-        4182 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Encodes mRFP1 under pLux promoter. Expresses a single guide RNA 
            (under Ara promoter), which targets a noncoding region of the 
            plasmid as part of the ADEPT system. Carries oriT and kan 
            resistance..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4182)
  AUTHORS   Son HI, Hamrick GS, Shende AR, Kim K, Yang K, Huang TJ, You L
  TITLE     Population-level amplification of gene regulation by programmable 
            gene transfer.
  JOURNAL   Nat Chem Biol. 2025 Jan 8. doi: 10.1038/s41589-024-01817-9.
  PUBMED    39779901
REFERENCE   2  (bases 1 to 4182)
  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"; journalName: "Nat Chem 
            Biol. 2025 Jan 8. doi: 10.1038/s41589-024-01817-9."
COMMENT     Sequence Label: ADEPT-pTarget-RFP
FEATURES             Location/Qualifiers
     source          1..4182
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      1..589
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     490..509
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     oriT            784..1074
                     /note="origin of transfer for the bacterial F plasmid 
                     (Frost et al., 1994)"
     primer_bind     1169..1186
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(1239..1285)
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     1438..1455
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     CDS             complement(1511..1528)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(1544..1573)
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     misc_RNA        complement(1594..1669)
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        complement(1690..1974)
                     /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     complement(1748..1767)
                     /label=pBAD Forward
                     /note="For vectors with E. coli araBAD promoter, forward 
                     primer"
     CDS             2135..2929
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(2189..2208)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2799..2818
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     terminator      2960..3054
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     terminator      complement(3164..3191)
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     terminator      complement(3207..3278)
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(3311..3988)
                     /codon_start=1
                     /product="monomeric derivative of DsRed (Campbell et al., 
                     2002)"
                     /label=mRFP1
                     /translation="MASSEDVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFEDGGV
                     VTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASTERMYPEDGALKGEIKM
                     RLKLKDGGHYDAEVKTTYMAKKPVQLPGAYKTDIKLDITSHNEDYTIVEQYERAEGRHS
                     TGA"
     RBS             3995..4006
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     promoter        complement(4016..4050)
                     /label=BBa_J23117
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.06"
ORIGIN
        1 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
       61 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
      121 cagcagagcg cagataccaa atactgtcct tctagtgtag ccgtagttag gccaccactt
      181 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
      241 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
      301 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
      361 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
      421 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
      481 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
      541 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctgtggaaaa acgccagcaa
      601 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
      661 gttatcccct gattctgtgg ataaccgtat taccgaggtt caccttagcc attagagatt
      721 accttttttg aacataaaat aaagttaatt tcgaaacact aaaattcata atgcaaacag
      781 ggacgcaccg ctagcagcgc ccctagcggt atcctataaa aaaacacacc gcgccgctag
      841 cagcacccct aatataaaat aatgtttttt ataaaaatag tcagtaccac ccctacaaaa
      901 cggtgtcggc gcgttgttgt agccgcgccg acaccgcttt tttaaatatc ataaagagag
      961 taagagaaac taatttttca taacactcta tttataaaga aaaatcagca aaaacttgtt
     1021 tttgcatggg gtgtggtgct tttggtggtg agaaccacca acctgttgag cctttttgtg
     1081 gagtgggtta aattatttac gaataaagtc accagaggtg gcctttgagt gagctgatac
     1141 cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg agcgaggaag cctgcagtct
     1201 agactcgagt aaggatccag ttcaccgaca aacaacagat aaaacgaaag gcccagtctt
     1261 tcgactgagc ctttcgtttt atttgatgcc tggcagttcc ctactctcgc atggggagac
     1321 cccacactac catcggcgct acggcgtttc acttctgagt tcggcatggg gtcaggtggg
     1381 accaccgcgc tactgccgcc aggcaaattc tgttttatca gaccgcttct gcgttctgat
     1441 ttaatctgta tcaggctgaa aatcttctct catccgccaa aacagccaag ctggagaccg
     1501 tttaaactca atgatgatga tgatgatggt cgacggcgct attcagatcc tcttctgaga
     1561 tgagtttttg ttcgggccca agcttcaaaa aaagcaccga ctcggtgcca ctttttcaag
     1621 ttgataacgg actagcctta ttttaacttg ctatttctag ctctaaaacc tgcccttcgc
     1681 ttgggacata tggagaaaca gtagagagtt gcgataaaaa gcgtcaggta ggatccgcta
     1741 atcttatgga taaaaatgct atggcatagc aaagtgtgac gccgtgcaaa taatcaatgt
     1801 ggacttttct gccgtgatta tagacacttt tgttacgcgt ttttgtcatg gctttggtcc
     1861 cgctttgtta cagaatgctt ttaataagcg gggttaccgg ttgggttagc gagaagagcc
     1921 agtaaaagac gcagtgacgg caatgtctga tgcaatatgg acaattggtt tcttggtttc
     1981 ttagacgtca gggacgtcgg aattgccagc tggggcgccc tctggtaagg ttgggaagcc
     2041 ctgcaaagta aactggatgg ctttcttgcc gccaaggatc tgatggcgca ggggatcaag
     2101 atctgatcaa gagacaggat gaggatcgtt tcgcatgatt gaacaagatg gattgcacgc
     2161 aggttctccg gccgcttggg tggagaggct attcggctat gactgggcac aacagacaat
     2221 cggctgctct gatgccgccg tgttccggct gtcagcgcag gggcgcccgg ttctttttgt
     2281 caagaccgac ctgtccggtg ccctgaatga actgcaggac gaggcagcgc ggctatcgtg
     2341 gctggccacg acgggcgttc cttgcgcagc tgtgctcgac gttgtcactg aagcgggaag
     2401 ggactggctg ctattgggcg aagtgccggg gcaggatctc ctgtcatctc accttgctcc
     2461 tgccgagaaa gtatccatca tggctgatgc aatgcggcgg ctgcatacgc ttgatccggc
     2521 tacctgccca ttcgaccacc aagcgaaaca tcgcatcgag cgagcacgta ctcggatgga
     2581 agccggtctt gtcgatcagg atgatctgga cgaagagcat caggggctcg cgccagccga
     2641 actgttcgcc aggctcaagg cgcgcatgcc cgacggcgag gatctcgtcg tgacccatgg
     2701 cgatgcctgc ttgccgaata tcatggtgga aaatggccgc ttttctggat tcatcgactg
     2761 tggccggctg ggtgtggcgg accgctatca ggacatagcg ttggctaccc gtgatattgc
     2821 tgaagagctt ggcggcgaat gggctgaccg cttcctcgtg ctttacggta tcgccgctcc
     2881 cgattcgcag cgcatcgcct tctatcgcct tcttgacgag ttcttctgag cgggactctg
     2941 gggttcgaga gctcgcttgg actcctgttg atagatccag taatgacctc agaactccat
     3001 ctggatttgt tcagaacgct cggttgccgc cgggcgtttt ttattggtga gaatccaagc
     3061 actagtaaca acttatatcg tatggggctg acttcaggtg ctaccgctag caacaccaga
     3121 acagcccgtt tgcgggcagc aaaacccgta ccctaggtat aaacgcagaa aggcccaccc
     3181 gaaggtgagc cagtgtgact ctagtagaga gcgttcaccg acaaacaaca gataaaacga
     3241 aaggcccagt ctttcgactg agcctttcgt tttatttgat gcctggagat ccttactcga
     3301 gtttggatcc ttaagcaccg gtggagtgac gaccttcagc acgttcgtac tgttcaacga
     3361 tggtgtagtc ttcgttgtgg gaggtgatgt ccagtttgat gtcggttttg taagcacccg
     3421 gcagctgaac cggtttttta gccatgtagg tggttttaac ttcagcgtcg tagtgaccac
     3481 cgtctttcag tttcagacgc attttgattt cacctttcag agcaccgtct tccgggtaca
     3541 tacgttcggt ggaagcttcc caacccatgg tttttttctg cataaccgga ccgtcggacg
     3601 ggaagttggt accacgcagt ttaactttgt agatgaactc accgtcttgc agggaggagt
     3661 cctgggtaac ggtaacaaca ccaccgtctt cgaagttcat aacacgttcc catttgaaac
     3721 cttccgggaa ggacagtttc aggtagtccg ggatgtcagc cgggtgttta acgtaagctt
     3781 tggaaccgta ctggaactgc ggggacagga tgtcccaagc gaacggcagc ggaccacctt
     3841 tggtaacttt cagtttagcg gtctgggtac cttcgtacgg acgaccttca ccttcacctt
     3901 cgatttcgaa ctcgtgaccg ttaacggaac cttccatacg aactttgaaa cgcatgaact
     3961 ctttgataac gtcttcgcta ctcgccatgg tacctttctc ctctttaatg aattcgctag
     4021 cacaatccct aggactgagc tagctgtcaa gtccggaggt atccggtgga acccaaagga
     4081 cgcctttggt aaccgcagga caccgcagga tacctgaggt cgcccgtggt ggcccatggt
     4141 caccataggt cacgaccccg tagaaaagat caaaggatct tc
//
