LOCUS       sequence-413787-        3322 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Empty C-terminus donor cassette. Integrate homology arms to target 
            the C-terminus insertion of a 3xFLAG-2A-Zeo cassette.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3322)
  AUTHORS   Philip R, Sharma A, Matellan L, Erpf AC, Hsu WH, Tkach JM, Wyatt 
            HDM, Pelletier L
  TITLE     qTAG: an adaptable plasmid scaffold for CRISPR-based endogenous 
            tagging.
  JOURNAL   EMBO J. 2024 Dec 12. doi: 10.1038/s44318-024-00337-5.
  PUBMED    39668248
REFERENCE   2  (bases 1 to 3322)
  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: "EMBO J. 
            2024 Dec 12. doi: 10.1038/s44318-024-00337-5."
COMMENT     Sequence Label: qTAG-C-3xFLAG-Zeo
FEATURES             Location/Qualifiers
     source          1..3322
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(29..51)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     151..170
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     364..386
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     378..395
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     379..395
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             444..515
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags"
                     /label=3xFLAG
                     /translation="DYKDDDDKDYKDDDDKDYKDDDDK"
     protein_bind    complement(537..570)
                     /label=lox71
                     /bound_moiety="Cre recombinase"
                     /note="Left element (LE) mutant of loxP (Araki et al., 
                     2010). Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             582..638
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             645..1016
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     protein_bind    complement(1023..1056)
                     /label=lox66
                     /bound_moiety="Cre recombinase"
                     /note="Right element (RE) mutant of loxP (Araki et al., 
                     2010). Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     primer_bind     complement(1101..1117)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1101..1117)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1114..1136)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1125..1141
                     /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(1149..1155,1156..1173,1174..1179))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1194..1215
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1332..1349)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1503..2091)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1583..1602)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2262..3053,3054..3122))
                     /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     2885..2904
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3123..3227)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3295..3313
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca
       61 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
      121 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
      181 accatatgcg gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc atcaggcgcc
      241 attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat
      301 tacgccagct ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta acgccagggt
      361 tttcccagtc acgacgttgt aaaacgacgg ccagtgaatt cggaggaagc ggaggatctg
      421 gcggtaccgg tggaggcgga tctgattaca aggatgacga cgataaggac tataaggacg
      481 atgatgacaa ggactacaaa gatgatgacg ataaaggagg tggcggatca acgcgttacc
      541 gttcgtatag catacattat acgaagttat ctggatccgg agctactaac ttcagcctgc
      601 tgaagcaggc cggagacgtg gaggagaacc ctggaccttc tagaatggcc aagttgacca
      661 gtgccgttcc ggtgctcacc gcgcgcgacg tcgccggagc ggtcgagttc tggaccgacc
      721 ggctcgggtt cagccgggac ttcgtggagg acgacttcgc cggtgtggtc cgggacgacg
      781 tgaccctgtt catcagcgcg gtccaggacc aggtggtgcc ggacaacacc ctggcctggg
      841 tgtgggtgcg cggcctggac gagctgtacg ccgagtggtc ggaggtcgtg tccacgaact
      901 tccgggacgc ctccgggccg gccatgaccg agatcggcga gcagccgtgg gggcgggagt
      961 tcgccctgcg cgacccggcc ggcaactgcg tgcacttcgt ggccgaggag caggaccccg
     1021 ggataacttc gtatagcata cattatacga acggtagcct cgagaccggt aagagcaagt
     1081 gaaagcttgg cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta tccgctcaca
     1141 attccacaca acatacgagc cggaagcata aagtgtaaag cctggggtgc ctaatgagtg
     1201 agctaactca cattaattgc gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg
     1261 tgccagctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc
     1321 tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta
     1381 tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag
     1441 aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg
     1501 tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg
     1561 tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg
     1621 cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga
     1681 agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc
     1741 tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt
     1801 aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact
     1861 ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg
     1921 cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt
     1981 accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt
     2041 ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct
     2101 ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg
     2161 gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt
     2221 aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt
     2281 gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc
     2341 gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg
     2401 cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc
     2461 gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg
     2521 gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca
     2581 ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga
     2641 tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct
     2701 ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg
     2761 cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca
     2821 accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata
     2881 cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct
     2941 tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact
     3001 cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa
     3061 acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc
     3121 atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga
     3181 tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga
     3241 aaagtgccac ctgacgtcta agaaaccatt attatcatga cattaaccta taaaaatagg
     3301 cgtatcacga ggccctttcg tc
//
