LOCUS       sequence-370749-        4023 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  PCR template for repair templates to tag a gene with a GFP and mIAA7
            AID degron using homology-directed repair for C. elegans..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4023)
  AUTHORS   Sepers JJ, Verstappen NHM, Vo AA, Ragle JM, Ruijtenberg S, Ward JD, 
            Boxem M
  TITLE     The mIAA7 degron improves auxin-mediated degradation in C. elegans.
  JOURNAL   G3 (Bethesda). 2022 Aug 27. pii: 6677660. doi: 
            10.1093/g3journal/jkac222.
  PUBMED    36029236
REFERENCE   2  (bases 1 to 4023)
  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: "G3 
            (Bethesda). 2022 Aug 27. pii: 6677660. doi: 
            10.1093/g3journal/jkac222."
COMMENT     Sequence Label: pJJS002
FEATURES             Location/Qualifiers
     source          1..4023
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(33..461)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(143..164)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     355..374
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     588..610
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     602..619
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     603..619
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     626..645
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        626..644
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             join(690..992,1046..1120,1172..1291,1343..1558)
                     /codon_start=1
                     /product="monomeric enhanced GFP, optimized with synthetic 
                     introns for expression in C. elegans"
                     /label=mEGFP
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTFTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLKE
                     FVTAAGITHGMDELYK"
     promoter        complement(1838..1856)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(1877..1893)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1877..1893)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1890..1912)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1901..1917
                     /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(1925..1931,1932..1949,1950..1955))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1970..1991
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2108..2125)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2279..2867)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2359..2378)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3038..3829,3830..3898))
                     /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     3661..3680
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3899..4003)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 cacctaaatt gtaagcgtta atattttgtt aaaattcgcg ttaaattttt gttaaatcag
       61 ctcatttttt aaccaatagg ccgaaatcgg caaaatccct tataaatcaa aagaatagac
      121 cgagataggg ttgagtgttg ttccagtttg gaacaagagt ccactattaa agaacgtgga
      181 ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat ggcccactac gtgaaccatc
      241 accctaatca agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga accctaaagg
      301 gagcccccga tttagagctt gacggggaaa gccggcgaac gtggcgagaa aggaagggaa
      361 gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc tgcgcgtaac
      421 caccacaccc gccgcgctta atgcgccgct acagggcgcg tcccattcgc cattcaggct
      481 gcgcaactgt tgggaagggc gatcggtgcg ggcctcttcg ctattacgcc agctggcgaa
      541 agggggatgt gctgcaaggc gattaagttg ggtaacgcca gggttttccc agtcacgacg
      601 ttgtaaaacg acggccagtg aattgtaata cgactcacta tagggcgaat tggggcagcg
      661 gtggcagtgg aggtaccggc ggaagcggta tgtccaaggg agaggaactc ttcaccggag
      721 tcgtcccaat cctcgtcgag ctcgacggag acgtcaacgg acacaagttc tccgtctcag
      781 gagagggaga gggagacgcc acctacggaa agctcaccct caagttcatc tgcaccaccg
      841 gaaagctccc agtcccatgg ccaaccctcg tcaccacctt cacttacgga gtccaatgct
      901 tctcccgtta cccagaccac atgaagcgtc acgacttctt caagtccgcc atgccagagg
      961 gatacgtcca agagcgtacc atcttcttca aggtaagttt aaacattaat taatactaac
     1021 taaccctgat tatttaaatt ttcaggacga cggaaactac aagacccgtg ccgaggtcaa
     1081 gttcgaggga gacaccctcg tcaaccgtat cgagctcaag gtaagtttaa acagttcggt
     1141 actaactaac catacatatt taaattttca gggaatcgac ttcaaggagg acggaaacat
     1201 cctcggacac aagctcgaat acaactacaa ctcccacaac gtctacatca tggccgacaa
     1261 gcaaaagaac ggaatcaagg tcaacttcaa ggtaagttta aacatgattt tactaactaa
     1321 ctaatctgat ttaaattttc agatccgtca caacatcgag gacggatctg tccaactcgc
     1381 cgaccactac caacaaaaca ccccaatcgg agacggacca gtcctcctcc cagacaacca
     1441 ctacctctcc acccaatccg ccctctccaa ggacccaaac gagaagcgtg accacatggt
     1501 cctcaaggag ttcgtcaccg ctgccggaat cacccacgga atggacgagc tctacaaggg
     1561 attctccgag accgtcgacc tcatgctcaa cctccaatcc aacaaggagg gatccgtcga
     1621 cctcaagaac gtctccgccg tcccaaagga gaagaccacc ctcaaggtaa gtttaaacat
     1681 atatatacta actaaccctg attatttaaa ttttcaggac ccatccaagc caccagccaa
     1741 ggcccaagtc gtcggatggc caccagtccg taactaccgt aagaacatga tgacccaaca
     1801 aaagacctcc tccgcggtgg agctccagct tttgttccct ttagtgaggg ttaatttcga
     1861 gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc
     1921 cacacaacat acgagccgga agcataaagt gtaaagcctg gggtgcctaa tgagtgagct
     1981 aactcacatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc
     2041 agctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt
     2101 ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag
     2161 ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca
     2221 tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt
     2281 tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc
     2341 gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct
     2401 ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg
     2461 tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca
     2521 agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact
     2581 atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta
     2641 acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta
     2701 actacggcta cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct
     2761 tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt
     2821 tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga
     2881 tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca
     2941 tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat
     3001 caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg
     3061 cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt
     3121 agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag
     3181 acccacgctc accggctcca gatttatcag caataaacca gccagccgga agggccgagc
     3241 gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag
     3301 ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca
     3361 tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa
     3421 ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga
     3481 tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata
     3541 attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca
     3601 agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg
     3661 ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg
     3721 ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg
     3781 cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag
     3841 gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac
     3901 tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca
     3961 tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag
     4021 tgc
//
