LOCUS       sequence-308930-        3618 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Crispr repair template: add insertion site specific homology arms by
            PCR before insertion.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3618)
  AUTHORS   Ashley GE, Duong T, Levenson MT, Martinez MAQ, Johnson LC, Hibshman 
            JD, Saeger HN, Palmisano NJ, Doonan R, Martinez-Mendez R, Davidson 
            BR, Zhang W, Ragle JM, Medwig-Kinney TN, Sirota SS, Goldstein B, 
            Matus DQ, Dickinson DJ, Reiner DJ, Ward JD
  TITLE     An expanded auxin-inducible degron toolkit for Caenorhabditis 
            elegans.
  JOURNAL   Genetics. 2021 Mar 31;217(3). pii: 6104563. doi: 
            10.1093/genetics/iyab006.
  PUBMED    33677541
REFERENCE   2  (bases 1 to 3618)
  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: "Genetics.";
            date: "2021-03-31"; volume: "217(3). pii: 6104563. doi"; pages: " 
            10.1093/genetics/iyab006"
COMMENT     Sequence Label: pJW2086
FEATURES             Location/Qualifiers
     source          1..3618
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      complement(159..186)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(278..364)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     413..435
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     427..444
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     428..444
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             1430..1450
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             1472..1603
                     /codon_start=1
                     /product="minimal degron from the auxin-responsive IAA17 
                     protein of Arabidopsis thaliana (Morawska and Ulrich, 
                     2013)"
                     /label=AID*
                     /note="acts as an auxin-inducible degron (AID) in the 
                     presence of TIR1"
                     /translation="PKDPAKPPAKAQVVGWPPVRSYRKNVMVSCQKSSGGPEAAAFVK"
     CDS             1631..1696
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     primer_bind     complement(1750..1769)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1751..1769)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1774..1790)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1774..1790)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1892..1911)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             1903..2712
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(1970..1989)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      2859..3447
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3348..3367
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3601..3618
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 gaagcggaag agcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa
       61 tgcagctggc acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat
      121 acgcgtaccg ctagccagga agagtttgta gaaacgcaaa aaggccatcc gtcaggatgg
      181 ccttctgctt agtttgatgc ctggcagttt atggcgggcg tcctgcccgc caccctccgg
      241 gccgttgctt cacaacgttc aaatccgctc ccggcggatt tgtcctactc aggagagcgt
      301 tcaccgacaa acaacagata aaacgaaagg cccagtcttc cgactgagcc tttcgtttta
      361 tttgatgcct ggcagttccc tactctcgcg ttaacgctag catggatgtt ttcccagtca
      421 cgacgttgta aaacgacggc cagtcttaag ctcgggcccc ggtggcggtg gatcgggagg
      481 aggaggttcg ggtggcggag gcagtggagg tggcggaagt ggcggcggtg gttcaggagg
      541 aggcggatcc atgagtaaag gagaagaatt gttcactgga gttgtcccaa tcctcgtcga
      601 gctcgacgga gacgtcaacg gacacaagtt ctccgtctcc ggagagggag agggagacgc
      661 cacctacgga aagctcaccc tcaagttcat ctgcaccacc ggaaagctcc cagtcccatg
      721 gccaaccctc gtcaccacct tctgctacgg agtccaatgc ttctcccgtt acccagacca
      781 catgaagcgt cacgacttct tcaagtccgc catgccagag ggatacgtcc aagagcgtac
      841 catcttcttc aaggtaagtt taaacatata tatactaact actgattatt taaattttca
      901 ggacgacgga aactacaaga cccgtgccga ggtcaagttc gagggagaca ccctcgtcaa
      961 ccgtatcgag ctcaaggtaa gtttaaacag ttcggtacta actaaccata catatttaaa
     1021 ttttcaggga atcgacttca aggaggacgg aaacatcctc ggacacaagc tcgagtacaa
     1081 ctacaactcc cacaacgtct acatcatggc cgacaagcaa aagaacggaa tcaaggtcaa
     1141 cttcaaggta agtttaaaca tgattttact aactaactaa tctgatttaa attttcagat
     1201 ccgtcacaac atcgaggacg gatccgtcca actcgccgac cactaccaac aaaacacccc
     1261 aatcggagac ggaccagtcc tcctcccaga caaccactac ctctccaccc aatccgccct
     1321 ctccaaggac ccaaacgaga agcgtgacca catggtcctc ctcgagttcg tcaccgccgc
     1381 cggaatcacc cacggaatgg acgagctcta caagggcggc agcggaggag agaatctgta
     1441 ctttcaatcc ggatccggag gtggcgggat gcctaaagat ccagccaaac ctccggccaa
     1501 ggcacaagtt gtgggatggc caccggtgag atcataccgg aagaacgtga tggtttcctg
     1561 ccaaaaatca agcggtggcc cggaggcggc ggcgttcgtg aagggtggtg gcggttctgg
     1621 ctcaggatct gactacaaag accatgacgg tgattataaa gatcatgata tcgattacaa
     1681 ggatgacgat gacaagggag catcgggagc ctcaggagca tcgtccggag gcgccatcca
     1741 gctgatatcc cctatagtga gtcgtattac atggtcatag ctgtttcctg gcagctctgg
     1801 cccgtgtctc aaaatctctg atgttacatt gcacaagata aaaatatatc atcatgaaca
     1861 ataaaactgt ctgcttacat aaacagtaat acaaggggtg ttatgagcca tattcaacgg
     1921 gaaacgtcga ggccgcgatt aaattccaac atggatgctg atttatatgg gtataaatgg
     1981 gctcgcgata atgtcgggca atcaggtgcg acaatctatc gcttgtatgg gaagcccgat
     2041 gcgccagagt tgtttctgaa acatggcaaa ggtagcgttg ccaatgatgt tacagatgag
     2101 atggtcagac taaactggct gacggaattt atgcctcttc cgaccatcaa gcattttatc
     2161 cgtactcctg atgatgcatg gttactcacc actgcgatcc ccggaaaaac agcattccag
     2221 gtattagaag aatatcctga ttcaggtgaa aatattgttg atgcgctggc agtgttcctg
     2281 cgccggttgc attcgattcc tgtttgtaat tgtcctttta acagcgatcg cgtatttcgt
     2341 ctcgctcagg cgcaatcacg aatgaataac ggtttggttg atgcgagtga ttttgatgac
     2401 gagcgtaatg gctggcctgt tgaacaagtc tggaaagaaa tgcataaact tttgccattc
     2461 tcaccggatt cagtcgtcac tcatggtgat ttctcacttg ataaccttat ttttgacgag
     2521 gggaaattaa taggttgtat tgatgttgga cgagtcggaa tcgcagaccg ataccaggat
     2581 cttgccatcc tatggaactg cctcggtgag ttttctcctt cattacagaa acggcttttt
     2641 caaaaatatg gtattgataa tcctgatatg aataaattgc agtttcattt gatgctcgat
     2701 gagtttttct aatcagaatt ggttaattgg ttgtaacact ggcagagcat tacgctgact
     2761 tgacgggacg gcgcaagctc atgaccaaaa tcccttaacg tgagttacgc gtcgttccac
     2821 tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc
     2881 gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat
     2941 caagagctac caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat
     3001 actgttcttc tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct
     3061 acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt
     3121 cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg
     3181 gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta
     3241 cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg
     3301 gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg
     3361 tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc
     3421 tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg
     3481 gccttttgct ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat
     3541 aaccgtatta ccgcctttga gtgagctgat accgctcgcc gcagccgaac gaccgagcgc
     3601 agcgagtcag tgagcgag
//
