LOCUS       sequence-370751-        4018 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  PCR template for repair templates to tag a gene with a mNeonGreen 
            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 4018)
  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 4018)
  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: pJJS005
FEATURES             Location/Qualifiers
     source          1..4018
                     /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"
     regulatory      1218..1227
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(1833..1851)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(1872..1888)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1872..1888)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1885..1907)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1896..1912
                     /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(1920..1926,1927..1944,1945..1950))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1965..1986
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2103..2120)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2274..2862)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2354..2373)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3033..3824,3825..3893))
                     /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     3656..3675
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3894..3998)
                     /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 catggtctcc aagggagagg aggacaacat ggcctccctc ccagccaccc
      961 acgagctcca catcttcgga tccatcaacg gagtcgactt cgacatggtc ggacaaggaa
     1021 ccggaaaccc aaacgacgga tacgaggagc tcaacctcaa gtccaccaag gtaagtttaa
     1081 acatatatat actaactaac cctgattatt taaattttca gggagacctc caattctccc
     1141 catggatcct cgtcccacac atcggatacg gattccacca atacctccca tacccagacg
     1201 gaatgtcccc attccaagcc gccatggtcg acggatccgg ataccaagtc caccgtacca
     1261 tgcaattcga ggacggagcc tccctcaccg tcaactaccg ttacacctac gagggatccc
     1321 acatcaaggt aagtttaaac agttcggtac taactaacca tacatattta aattttcagg
     1381 gagaggccca agtcaaggga accggattcc cagccgacgg accagtcatg accaactccc
     1441 tcaccgccgc cgactggtgc cgttccaaga agacctaccc aaacgacaag gtaagtttaa
     1501 acatgatttt actaactaac taatctgatt taaattttca gaccatcatc tccaccttca
     1561 agtggtccta caccaccgga aacggaaagc gttaccgttc caccgcccgt accacctaca
     1621 ccttcgccaa gccaatggcc gccaactacc tcaagaacca accaatgtac gtcttccgta
     1681 agaccgagct caagcactcc aagaccgagc tcaacttcaa ggagtggcaa aaggccttca
     1741 ccgacgtcat gggaatggac gagctctaca agggcagcgg tggcagtgga ggtaccggcg
     1801 gaagcggtgc ggtggagctc cagcttttgt tccctttagt gagggttaat ttcgagcttg
     1861 gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt atccgctcac aattccacac
     1921 aacatacgag ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt gagctaactc
     1981 acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg
     2041 cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctcttccgct
     2101 tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac
     2161 tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga
     2221 gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat
     2281 aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac
     2341 ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct
     2401 gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg
     2461 ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg
     2521 ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt
     2581 cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg
     2641 attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac
     2701 ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga
     2761 aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt
     2821 gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt
     2881 tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga
     2941 ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc
     3001 taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct
     3061 atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata
     3121 actacgatac gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca
     3181 cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga
     3241 agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga
     3301 gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg
     3361 gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga
     3421 gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt
     3481 gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct
     3541 cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca
     3601 ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat
     3661 accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga
     3721 aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc
     3781 aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg
     3841 caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc
     3901 ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt
     3961 gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgc
//
