LOCUS       sequence-370743-        4026 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 4026)
  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 4026)
  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: pJJS001
FEATURES             Location/Qualifiers
     source          1..4026
                     /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(912..1214,1268..1342,1394..1513,1565..1780)
                     /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(1841..1859)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(1880..1896)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1880..1896)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1893..1915)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1904..1920
                     /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(1928..1934,1935..1952,1953..1958))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1973..1994
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2111..2128)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2282..2870)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2362..2381)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3041..3832,3833..3901))
                     /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     3664..3683
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3902..4006)
                     /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 tggatgggat
      661 tctccgagac cgtcgacctc atgctcaacc tccaatccaa caaggaggga tccgtcgacc
      721 tcaagaacgt ctccgccgtc ccaaaggaga agaccaccct caaggtaagt ttaaacatat
      781 atatactaac taaccctgat tatttaaatt ttcaggaccc atccaagcca ccagccaagg
      841 cccaagtcgt cggatggcca ccagtccgta actaccgtaa gaacatgatg acccaacaaa
      901 agacctcctc catgtccaag ggagaggaac tcttcaccgg agtcgtccca atcctcgtcg
      961 agctcgacgg agacgtcaac ggacacaagt tctccgtctc aggagaggga gagggagacg
     1021 ccacctacgg aaagctcacc ctcaagttca tctgcaccac cggaaagctc ccagtcccat
     1081 ggccaaccct cgtcaccacc ttcacttacg gagtccaatg cttctcccgt tacccagacc
     1141 acatgaagcg tcacgacttc ttcaagtccg ccatgccaga gggatacgtc caagagcgta
     1201 ccatcttctt caaggtaagt ttaaacatta attaatacta actaaccctg attatttaaa
     1261 ttttcaggac gacggaaact acaagacccg tgccgaggtc aagttcgagg gagacaccct
     1321 cgtcaaccgt atcgagctca aggtaagttt aaacagttcg gtactaacta accatacata
     1381 tttaaatttt cagggaatcg acttcaagga ggacggaaac atcctcggac acaagctcga
     1441 atacaactac aactcccaca acgtctacat catggccgac aagcaaaaga acggaatcaa
     1501 ggtcaacttc aaggtaagtt taaacatgat tttactaact aactaatctg atttaaattt
     1561 tcagatccgt cacaacatcg aggacggatc tgtccaactc gccgaccact accaacaaaa
     1621 caccccaatc ggagacggac cagtcctcct cccagacaac cactacctct ccacccaatc
     1681 cgccctctcc aaggacccaa acgagaagcg tgaccacatg gtcctcaagg agttcgtcac
     1741 cgctgccgga atcacccacg gaatggacga gctctacaag ggcagcggtg gcagtggagg
     1801 taccggcgga agcggtgcgg tggagctcca gcttttgttc cctttagtga gggttaattt
     1861 cgagcttggc gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa
     1921 ttccacacaa catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga
     1981 gctaactcac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt
     2041 gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct
     2101 cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat
     2161 cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga
     2221 acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt
     2281 ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt
     2341 ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc
     2401 gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa
     2461 gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct
     2521 ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta
     2581 actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg
     2641 gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc
     2701 ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta
     2761 ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg
     2821 gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt
     2881 tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg
     2941 tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta
     3001 aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg
     3061 aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg
     3121 tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc
     3181 gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg
     3241 agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg
     3301 aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag
     3361 gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat
     3421 caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc
     3481 cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc
     3541 ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa
     3601 ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac
     3661 gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt
     3721 cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc
     3781 gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa
     3841 caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca
     3901 tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat
     3961 acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa
     4021 aagtgc
//
