LOCUS       sequence-274806-        3398 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasmid for PCR-generated donor DNA for rapid tagging of proteins 
            with tagRFP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3398)
  AUTHORS   Paix A, Rasoloson D, Folkmann A, Seydoux G
  TITLE     Rapid Tagging of Human Proteins with Fluorescent Reporters by Genome
            Engineering using Double-Stranded DNA Donors.
  JOURNAL   Curr Protoc Mol Biol. 2019 Dec;129(1):e102. doi: 10.1002/cpmb.102.
  PUBMED    31710422
REFERENCE   2  (bases 1 to 3398)
  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: "Curr Protoc
            Mol Biol."; date: "2019-12"; volume: "129(1):e102. doi"; pages: " 
            10.1002/cpmb.102"
COMMENT     Sequence Label: pAP1680
FEATURES             Location/Qualifiers
     source          1..3398
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1..17
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(799..815)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(799..815)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(812..834)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    823..839
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(847..853,854..871,872..877))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    892..913
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1030..1047)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1201..1789)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1281..1300)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1960..2751,2752..2820))
                     /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     2583..2602
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2821..2925)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     2993..3011
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(3049..3071)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     3171..3190
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     join(3384..3398,1..8)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     join(3398,1..17)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
ORIGIN
        1 gtaaaacgac ggccagtgaa ttcgagctcg gtacccgggg atcgtctcta aaggggaaga
       61 actcatcaag gagaatatgc acatgaagct ctatatggaa ggtacagtaa acaatcacca
      121 cttcaaatgc acttctgaag gtgaaggaaa gccctacgag ggtacacaga cgatgagaat
      181 taaagtcgtt gagggaggcc cgctgccctt tgcttttgac atccttgcaa cctcatttat
      241 gtatggcagc cgaacattca tcaatcatac acaggggatt ccagattttt tcaaacagtc
      301 ctttccggag ggctttacat gggagcgagt aacgacctat gaagatgggg gagttttgac
      361 cgccacgcaa gatacgtcac tccaagacgg ctgcctgatt tacaatgtaa aaataagagg
      421 cgttaatttt ccgtctaatg gcccggtaat gcagaaaaag accctgggtt gggaggccaa
      481 cacggaaatg ctttatccgg cagacggtgg cttggagggg cgctctgaca tggcactgaa
      541 gttggtcggc ggaggtcacc ttatttgcaa cttcaaaact acgtaccgct caaagaagcc
      601 agccaaaaac cttaaaatgc ccggggttta ttatgtagac caccgcctcg aacggatcaa
      661 ggaagctgat aaagaaacgt atgtagagca acatgaagtc gcagtggcaa ggtactgtga
      721 tctccccagt aagctggggc acaaactgaa tgatcctcta gagtcgacct gcaggcatgc
      781 aagcttggcg taatcatggt catagctgtt tcctgtgtga aattgttatc cgctcacaat
      841 tccacacaac atacgagccg gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag
      901 ctaactcaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg
      961 ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc
     1021 ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc
     1081 agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa
     1141 catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt
     1201 tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg
     1261 gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg
     1321 ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag
     1381 cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc
     1441 caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa
     1501 ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg
     1561 taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc
     1621 taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac
     1681 cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg
     1741 tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt
     1801 gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt
     1861 catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa
     1921 atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga
     1981 ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt
     2041 gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg
     2101 agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga
     2161 gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga
     2221 agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg
     2281 catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc
     2341 aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc
     2401 gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca
     2461 taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac
     2521 caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg
     2581 ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc
     2641 ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg
     2701 tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac
     2761 aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat
     2821 actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata
     2881 catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa
     2941 agtgccacct gacgtctaag aaaccattat tatcatgaca ttaacctata aaaataggcg
     3001 tatcacgagg ccctttcgtc tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat
     3061 gcagctcccg gagacggtca cagcttgtct gtaagcggat gccgggagca gacaagcccg
     3121 tcagggcgcg tcagcgggtg ttggcgggtg tcggggctgg cttaactatg cggcatcaga
     3181 gcagattgta ctgagagtgc accatatgcg gtgtgaaata ccgcacagat gcgtaaggag
     3241 aaaataccgc atcaggcgcc attcgccatt caggctgcgc aactgttggg aagggcgatc
     3301 ggtgcgggcc tcttcgctat tacgccagct ggcgaaaggg ggatgtgctg caaggcgatt
     3361 aagttgggta acgccagggt tttcccagtc acgacgtt
//
