LOCUS       sequence-320618-        3480 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 3480)
  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 3480)
  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: pJW2172
FEATURES             Location/Qualifiers
     source          1..3480
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     51..73
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     65..82
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     66..82
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     regulatory      495..504
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1065..1085
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             1131..1196
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     primer_bind     complement(1250..1269)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1251..1269)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1274..1290)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1274..1290)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1392..1411)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             1403..2212
                     /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(1470..1489)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      2359..2947
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2848..2867
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3101..3118
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(3277..3304)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(join(3396..3480,1..2))
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
ORIGIN
        1 tgatgcctgg cagttcccta ctctcgcgtt aacgctagca tggatgtttt cccagtcacg
       61 acgttgtaaa acgacggcca gtcttaagct cgggccccgg tggcggtgga tcgggaggag
      121 gaggttcggg tggcggaggc agtggaggtg gcggaagtgg cggcggtggt tcaggaggag
      181 gcggatccat ggtctccaag ggagaggagg acaacatggc ctccctccca gccacccacg
      241 agctccacat cttcggatcc atcaacggag tcgacttcga catggtcgga caaggaaccg
      301 gaaacccaaa cgacggatac gaggagctca acctcaagtc caccaaggta agtttaaaca
      361 tatatatact aactaaccct gattatttaa attttcaggg agacctccaa ttctccccat
      421 ggatcctcgt cccacacatc ggatacggat tccaccaata cctcccatac ccagacggaa
      481 tgtccccatt ccaagccgcc atggtcgacg gatccggata ccaagtccac cgtaccatgc
      541 aattcgagga cggagcctcc ctcaccgtca actaccgtta cacctacgag ggatcccaca
      601 tcaaggtaag tttaaacagt tcggtactaa ctaaccatac atatttaaat tttcagggag
      661 aggcccaagt caagggaacc ggattcccag ccgacggacc agtcatgacc aactccctca
      721 ccgccgccga ctggtgccgt tccaagaaga cctacccaaa cgacaaggta agtttaaaca
      781 tgattttact aactaactaa tctgatttaa attttcagac catcatctcc accttcaagt
      841 ggtcctacac caccggaaac ggaaagcgtt accgttccac cgcccgtacc acctacacct
      901 tcgccaagcc aatggccgcc aactacctca agaaccaacc aatgtacgtc ttccgtaaga
      961 ccgagctcaa gcactccaag accgagctca acttcaagga gtggcaaaag gccttcaccg
     1021 acgtcatggg aatggacgag ctctacaagg gcggcagcgg aggagagaat ctgtactttc
     1081 aatccggatc cggaggtggc gggggtggtg gcggttctgg ctcaggatct gactacaaag
     1141 accatgacgg tgattataaa gatcatgata tcgattacaa ggatgacgat gacaagggag
     1201 catcgggagc ctcaggagca tcgtccggag gcgccatcca gctgatatcc cctatagtga
     1261 gtcgtattac atggtcatag ctgtttcctg gcagctctgg cccgtgtctc aaaatctctg
     1321 atgttacatt gcacaagata aaaatatatc atcatgaaca ataaaactgt ctgcttacat
     1381 aaacagtaat acaaggggtg ttatgagcca tattcaacgg gaaacgtcga ggccgcgatt
     1441 aaattccaac atggatgctg atttatatgg gtataaatgg gctcgcgata atgtcgggca
     1501 atcaggtgcg acaatctatc gcttgtatgg gaagcccgat gcgccagagt tgtttctgaa
     1561 acatggcaaa ggtagcgttg ccaatgatgt tacagatgag atggtcagac taaactggct
     1621 gacggaattt atgcctcttc cgaccatcaa gcattttatc cgtactcctg atgatgcatg
     1681 gttactcacc actgcgatcc ccggaaaaac agcattccag gtattagaag aatatcctga
     1741 ttcaggtgaa aatattgttg atgcgctggc agtgttcctg cgccggttgc attcgattcc
     1801 tgtttgtaat tgtcctttta acagcgatcg cgtatttcgt ctcgctcagg cgcaatcacg
     1861 aatgaataac ggtttggttg atgcgagtga ttttgatgac gagcgtaatg gctggcctgt
     1921 tgaacaagtc tggaaagaaa tgcataaact tttgccattc tcaccggatt cagtcgtcac
     1981 tcatggtgat ttctcacttg ataaccttat ttttgacgag gggaaattaa taggttgtat
     2041 tgatgttgga cgagtcggaa tcgcagaccg ataccaggat cttgccatcc tatggaactg
     2101 cctcggtgag ttttctcctt cattacagaa acggcttttt caaaaatatg gtattgataa
     2161 tcctgatatg aataaattgc agtttcattt gatgctcgat gagtttttct aatcagaatt
     2221 ggttaattgg ttgtaacact ggcagagcat tacgctgact tgacgggacg gcgcaagctc
     2281 atgaccaaaa tcccttaacg tgagttacgc gtcgttccac tgagcgtcag accccgtaga
     2341 aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac
     2401 aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt
     2461 tccgaaggta actggcttca gcagagcgca gataccaaat actgttcttc tagtgtagcc
     2521 gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat
     2581 cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag
     2641 acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc
     2701 cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag
     2761 cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac
     2821 aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg
     2881 gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct
     2941 atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc
     3001 tcacatgttc tttcctgcgt tatcccctga ttctgtggat aaccgtatta ccgcctttga
     3061 gtgagctgat accgctcgcc gcagccgaac gaccgagcgc agcgagtcag tgagcgagga
     3121 agcggaagag cgcccaatac gcaaaccgcc tctccccgcg cgttggccga ttcattaatg
     3181 cagctggcac gacaggtttc ccgactggaa agcgggcagt gagcgcaacg caattaatac
     3241 gcgtaccgct agccaggaag agtttgtaga aacgcaaaaa ggccatccgt caggatggcc
     3301 ttctgcttag tttgatgcct ggcagtttat ggcgggcgtc ctgcccgcca ccctccgggc
     3361 cgttgcttca caacgttcaa atccgctccc ggcggatttg tcctactcag gagagcgttc
     3421 accgacaaac aacagataaa acgaaaggcc cagtcttccg actgagcctt tcgttttatt
//
