LOCUS       sequence-127823-        3638 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses myc::TEV::3Xflag::eGFP::3Xflag::TEV::myc in bacteria.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3638)
  AUTHORS   Paix A, Folkmann A, Rasoloson D, Seydoux G
  TITLE     High Efficiency, Homology-Directed Genome Editing in Caenorhabditis 
            elegans Using CRISPR-Cas9 Ribonucleoprotein Complexes.
  JOURNAL   Genetics. 2015 Sep;201(1):47-54. doi: 10.1534/genetics.115.179382. 
            Epub 2015 Jul 17.
  PUBMED    26187122
REFERENCE   2  (bases 1 to 3638)
  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: 
            "10.1534/genetics.115.179382"; journalName: "Genetics"; date: 
            "2015-09"; volume: "201"; issue: "1"; pages: "47-54"
COMMENT     Sequence Label: AP625-1
FEATURES             Location/Qualifiers
     source          1..3638
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    106..127
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(142..147,148..165,166..172)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    180..196
                     /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)."
     primer_bind     185..207
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     204..220
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     204..220
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             complement(268..297)
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             complement(298..318)
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             complement(319..384)
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             complement(join(385..1095,1096..1098))
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(425..445)
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     primer_bind     1040..1068
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     CDS             complement(1099..1164)
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             complement(1165..1185)
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             complement(1186..1215)
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     primer_bind     complement(1242..1259)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1242..1258)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1251..1273)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(1467..1486)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     1586..1608
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(1646..1664)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1732..1836
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1837..1905,1906..2697)
                     /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     complement(2055..2074)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2868..3456
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3357..3376
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3610..3627
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca
       61 cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaatg tgagttagct
      121 cactcattag gcaccccagg ctttacactt tatgcttccg gctcgtatgt tgtgtggaat
      181 tgtgagcgga taacaatttc acacaggaaa cagctatgac catgattacg ccaagcttgc
      241 atgcctgcag gtcgactcta gaggatccag atcctcttct gatatcagtt tttgttctcc
      301 ttggaagtag aggttctcct tgtcatcgtc atccttgtaa tcgatatcat gatctttata
      361 atcaccgtca tggtctttgt agtccttgta cagctcgtcc atgccgagtg taatcccagc
      421 agctgttaca aactcaagaa ggaccatgtg gtctctcttt tcgttgggat ctttcgaaag
      481 ggcagattgt gtggacaggt aatggttgtc tggtaaaagg acagggccat cgccaattgg
      541 agtattttgt tgataatggt ctgctagttg aacgcttcca tcttcaatgt tgtgtctaat
      601 tttgaagtta actttgattc cattcttttg tttgtctgcc atgatgtata cattgtgtga
      661 gttatagttg tattccaatt tgtgtccaag aatgtttcca tcttctttaa aatcaatacc
      721 ttttaactcg attctattaa caagggtatc accttcaaac ttgacttcag cacgtgtctt
      781 gtagttcccg tcatctttga aaaatatagt tctttcctgt acataacctt cgggcatggc
      841 actcttgaaa aagtcatgct gcttcatgtg gtcggggtac cggctgaagc actgcacgcc
      901 gtaggtcaga gtagtgacaa gtgttggcca tggaacaggt agttttccag tagtgcaaat
      961 aaatttaagg gtaagttttc cgtatgttgc atcaccttca ccctctccac tgacagaaaa
     1021 tttgtgccca ttaacatcac catctaattc aacaagaatt gggacaactc cagtgaaaag
     1081 ttcttctcct ttactcacct tgtcgtcgtc gtccttgtag tcgatgtcgt ggtccttgta
     1141 gtctccgtcg tggtccttgt agtcaccctg aaaatataaa ttttcgaggt cctcctcgga
     1201 gatgagcttt tgctcgatcc ccgggtaccg agctcgaatt cactggccgt cgttttacaa
     1261 cgtcgtgact gggaaaaccc tggcgttacc caacttaatc gccttgcagc acatccccct
     1321 ttcgccagct ggcgtaatag cgaagaggcc cgcaccgatc gcccttccca acagttgcgc
     1381 agcctgaatg gcgaatggcg cctgatgcgg tattttctcc ttacgcatct gtgcggtatt
     1441 tcacaccgca tatggtgcac tctcagtaca atctgctctg atgccgcata gttaagccag
     1501 ccccgacacc cgccaacacc cgctgacgcg ccctgacggg cttgtctgct cccggcatcc
     1561 gcttacagac aagctgtgac cgtctccggg agctgcatgt gtcagaggtt ttcaccgtca
     1621 tcaccgaaac gcgcgagacg aaagggcctc gtgatacgcc tatttttata ggttaatgtc
     1681 atgataataa tggtttctta gacgtcaggt ggcacttttc ggggaaatgt gcgcggaacc
     1741 cctatttgtt tatttttcta aatacattca aatatgtatc cgctcatgag acaataaccc
     1801 tgataaatgc ttcaataata ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc
     1861 gcccttattc ccttttttgc ggcattttgc cttcctgttt ttgctcaccc agaaacgctg
     1921 gtgaaagtaa aagatgctga agatcagttg ggtgcacgag tgggttacat cgaactggat
     1981 ctcaacagcg gtaagatcct tgagagtttt cgccccgaag aacgttttcc aatgatgagc
     2041 acttttaaag ttctgctatg tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa
     2101 ctcggtcgcc gcatacacta ttctcagaat gacttggttg agtactcacc agtcacagaa
     2161 aagcatctta cggatggcat gacagtaaga gaattatgca gtgctgccat aaccatgagt
     2221 gataacactg cggccaactt acttctgaca acgatcggag gaccgaagga gctaaccgct
     2281 tttttgcaca acatggggga tcatgtaact cgccttgatc gttgggaacc ggagctgaat
     2341 gaagccatac caaacgacga gcgtgacacc acgatgcctg tagcaatggc aacaacgttg
     2401 cgcaaactat taactggcga actacttact ctagcttccc ggcaacaatt aatagactgg
     2461 atggaggcgg ataaagttgc aggaccactt ctgcgctcgg cccttccggc tggctggttt
     2521 attgctgata aatctggagc cggtgagcgt gggtctcgcg gtatcattgc agcactgggg
     2581 ccagatggta agccctcccg tatcgtagtt atctacacga cggggagtca ggcaactatg
     2641 gatgaacgaa atagacagat cgctgagata ggtgcctcac tgattaagca ttggtaactg
     2701 tcagaccaag tttactcata tatactttag attgatttaa aacttcattt ttaatttaaa
     2761 aggatctagg tgaagatcct ttttgataat ctcatgacca aaatccctta acgtgagttt
     2821 tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt
     2881 tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt
     2941 ttgccggatc aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag
     3001 ataccaaata ctgttcttct agtgtagccg tagttaggcc accacttcaa gaactctgta
     3061 gcaccgccta catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat
     3121 aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg
     3181 ggctgaacgg ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg
     3241 agatacctac agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac
     3301 aggtatccgg taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga
     3361 aacgcctggt atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt
     3421 ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta
     3481 cggttcctgg ccttttgctg gccttttgct cacatgttct ttcctgcgtt atcccctgat
     3541 tctgtggata accgtattac cgcctttgag tgagctgata ccgctcgccg cagccgaacg
     3601 accgagcgca gcgagtcagt gagcgaggaa gcggaaga
//
