LOCUS       sequence-178587-        8272 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Lox2272-flanked SEC donor for the SapTrap cloning system.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8272)
  AUTHORS   Dickinson D, Slabodnick M, Chen A, Goldstein B
  TITLE     SapTrap assembly of repair templates for Cas9-triggered homologous 
            recombination with a self-excising cassette.
  JOURNAL   MicroPubl Biol. 2018 May 1;2018:10.17912/W2KT0N. doi: 
            10.17912/W2KT0N.
  PUBMED    32550377
REFERENCE   2  (bases 1 to 8272)
  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: "MicroPubl 
            Biol."; date: "2018-05-1"; volume: "2018:10.17912/W2KT0N. doi"; 
            pages: " 10.17912/W2KT0N"
COMMENT     Sequence Label: pDD382
FEATURES             Location/Qualifiers
     source          1..8272
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     92..109
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(268..295)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(387..473)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     522..544
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     536..553
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     537..553
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(624..747)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     protein_bind    complement(770..803)
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     CDS             813..842
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     terminator      complement(2659..2982)
                     /gene="tbb-2"
                     /label=tbb-2 terminator
                     /note="transcription terminator for the C. elegans tubulin 
                     beta-2 gene"
     protein_bind    4900..4924
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             4927..5952
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     CDS             4927..5952
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     protein_bind    complement(5953..5977)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(6330..6363)
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     protein_bind    complement(6381..6476)
                     /gene="mutant version of attL"
                     /label=attL3
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(6513..6532)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(6514..6532)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(6537..6553)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6537..6553)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6655..6674)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             6666..7475
                     /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(6733..6752)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      7622..8210
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8111..8130
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga
       61 taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga
      121 gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca
      181 cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaata cgcgtaccgc
      241 tagccaggaa gagtttgtag aaacgcaaaa aggccatccg tcaggatggc cttctgctta
      301 gtttgatgcc tggcagttta tggcgggcgt cctgcccgcc accctccggg ccgttgcttc
      361 acaacgttca aatccgctcc cggcggattt gtcctactca ggagagcgtt caccgacaaa
      421 caacagataa aacgaaaggc ccagtcttcc gactgagcct ttcgttttat ttgatgcctg
      481 gcagttccct actctcgcgt taacgctagc atggatgttt tcccagtcac gacgttgtaa
      541 aacgacggcc agtcttaagc tcgggccctg cagctctaga gctcgaattc tacaggtcac
      601 taataccatc taagtagttg gttcatagtg actgcatatg ttgtgtttta cagtattatg
      661 tagtctgttt tttatgcaaa atctaattta atatattgat atttatatca ttttacgttt
      721 ctcgttcaac tttcttgtac aaagtgggct cttcgaaggt aagtttaaaa taacttcgta
      781 taggatactt tatacgaagt tattttcagg gagagcaaaa gcttatctcc gaggaggacc
      841 tctaaggatg atcgacgcca acgtcgttga attttcaaat tttaaatact gaatatttgt
      901 tttttttcct attatttatt tattctcttt gtgttttttt tcttgctttc taaaaaatta
      961 attcaatcca aatctaaaca tttttttttc tctttccgtc tcccaattcg tattccgctc
     1021 ctctcatctg aacacaatgt gcaagtttat ttatcttctc gctttcattt cattaggacg
     1081 tggggggaat tggtggaagg gggaaacaca caaaaggatg atggaaatga aataaggaca
     1141 cacaatatgc aacaacattc aattcagaaa tatggaggaa ggtttaaaag aaaacataaa
     1201 aatatataga ggaggaagga aactagctgc cctcgattgt ctatcttcca ttctcaccaa
     1261 gtttcaacaa ctttattcag tcattcttct ctggttcatt tcccaaccaa tcttcccgaa
     1321 taaagcccat ctgtcctttc ggccacattc ctcacctaca tccgtactcc ttatctcccg
     1381 ttcttatcca gtcttttatc gtctctgcgt ctcgcacact gtctcgtcgg taatgtaaaa
     1441 aaaaaacaaa aaaaacttga aaacttgcgg aaagatttcc taataaaagg agacgaatcc
     1501 ggtaggaaat cgacaagtct ctgcttctcg aagactaggt gagttacatt tctttggccg
     1561 aattattttg aaatcctaaa tttgataatt tcagagatgt ctgtaaaact tgcgtgttat
     1621 gtgacggctt cggtcactgt cgccactctt atggtatgct tcatgaccat gtcaaccatc
     1681 tactcggagg ttgatggatt cagagagaag cttgataccg agatgaatgt gttcagagtg
     1741 agttaaatta tattttgaat tttaaataat ttttaatttt ctagcaatct accaatggat
     1801 tgtggaagga catagttgtc atcggaagat ctagcaagcg tgtccgttgt caatatgaag
     1861 agaccaacgc taccccaact ccacatgctg atggatcccc atctgctcca ccaggtcaac
     1921 caccagcagt tccaccagtc ttcaaccagc caaagactcc aaatggagcc aatggaaatg
     1981 gaccaacctg caactgcaat gctgataaca agtgcccagc tggaccatcc ggaccaaagg
     2041 gagttccagg agttccagga ctcgacggag ttccaggact tgacggtgtt ccaggagttg
     2101 gagctgatga tatcgctcca caacgcgagt ctgtcggatg cttcacttgc ccacaaggac
     2161 cagttggacc accaggagct cttggaagac caggaccacg tggacttcca ggaccaagag
     2221 gacaaaatgg aaacccagga agagatggac aaccaggaca tccaggagag caaggatcat
     2281 ccggtcaaat cggaaagatc ggagagccag gaccaccagg agagaaggga cgcgacgccg
     2341 agcatccaat cggaagacca ggaccaaagg gaccaagagg agatcaagga ccaacaggac
     2401 cagctggaca gaacggtctt cacggaccac caggagagcc aggaaccgtt ggaccagaag
     2461 gaccatctgg aaagcaagga cgtcaaggac cagacggaac ccagggagag actggaccag
     2521 acggaagacc aggaaaggat gccgagtact gccagtgccc agacaagtct ccaccatcag
     2581 aggctgtcaa cgccaaccgt ggatacagaa atatctaaat tgttggtgtt ttctaataaa
     2641 atatttgaga ttcagtgaga cttttttctt ggcggcacaa taaagtttcc tctttttgtt
     2701 gggtcactca acgaacacaa tacaaatatt tgagcgctcg catttttctc aacctttcgg
     2761 gtttaaagca cttcaaaatg gaattgaaga gagattgtga aagtacatca gacagaatca
     2821 cattttgata gcattcactt cactcagatg caaacttttc agtgactata aaaagtcata
     2881 aaataataac attaaaaaag caaataaatt agcgagagaa ttttttgaca aaaagaaaga
     2941 agagtgatag agaagaaggg aatgcttgaa aggattttgc atttatctta atcaccgtct
     3001 tcgagaagac gaaccatggc gccagtttca gaatccaagt ttctaatata gttcatcaca
     3061 atgttgacgt tggtccaacc tccggcttgc ataatctctg ggatggaaac tccagcgcga
     3121 gccatgtcac gggcagctcc gacacgggcg gaatgtccag accaagccaa gtaacgttgt
     3181 ccggaatcat ccttggctcc gtagatgaga cggtgggtag cttcgaagat tcctgaaaat
     3241 ttattttcag cttattctta gctaatatgt tgtttaaact taccttcaag agctcgagtc
     3301 gagagttgtg atgttgcact cggggcagca actccgttct tgcgaacacg acagaacagg
     3361 tagttgtttg gatcatcggc gactccggaa accgaaatcc agcgctcaac gagcttggta
     3421 actccaaggg agagcgcttt ttcgactccg gcagtcgaga ccagcgtttt tgtacgtccg
     3481 atgtggatga gcattcttcc tccgtctgtg cgactaatat ccttgacgcg aatgcgggca
     3541 atttcggcga tacggagaag tgtgttgtaa gcaattccga ggaaggcgag gtttcggatg
     3601 tcctggcaac gatcggaatt ttccataaga gaacggacct gatcgaagtc ggttctgaaa
     3661 atttcgtcaa gtcttaggat tgaatacatt aatatttaaa cttacctttc gaaggcgagg
     3721 gcttgcttgg ctctctctcc agcatcaaca ttttccttgc ggatacgtct cattacgaga
     3781 gatacggcat tagaatctga tgggcgtgga agtccagatc ggcgatggag catattgagc
     3841 tgtccgagat gttgttggat cgtcttcacc gccaatccac gagcttggag gtagagcaag
     3901 tagtcacgaa cgtcttctgg ctcggctggg aaccacttgc ggttgttaag cttgcaccag
     3961 gcagcccaag aacggcacac tgagagaagc atcttccagg tgtgctcgga gaaggcttgt
     4021 ctgtcacgga acatatccat gaggttcttg cgaacttcat cacttgtcgc gtcaactgga
     4081 agtgctggaa gattttgatg aacagtgaga agatttgaca tattttcgaa gttttttaga
     4141 tgcactagaa caaagcgtgt tggcttcctc tgagcccgct ttccttatat acccgcattc
     4201 tgcagcctta cagaatgttc tagaaggtcc tagatgcatt cgtttgaaaa tactcccggt
     4261 gggtgcaaag agacgcagac ggaaaatgta tctgggtctc tttattgtgt acactacttt
     4321 tccatgtacc gaatgtgagt cgccctcctt ttgcaacaag cagctcgaat gttctagaaa
     4381 aaggtggaaa atagtataaa taccgttgaa aataaatacc gaacaacatt tgctctaatt
     4441 gtgaaattag aaatcttcaa actataatca tgtcacttta ccactatttc cgtccagctc
     4501 aacgttccgt ttttggtgat catttttgct ttcgtcgtaa atctacacac gcgtctcttc
     4561 cgtgcgagag tccaagccag cagccaaatt cgttgactga gtattcaacg tttatacgtt
     4621 gtcggcaacg agaaatagga aaatgcatcg ggaaatgttc ttttttcgat tttttccaag
     4681 gttttgacaa attttaccac gaattttgct atgttttcaa ttaaaaaata tgttattcaa
     4741 ctgtttctat gaggaaaata aggctttgca tgtaattttc ttattcagca taatttttaa
     4801 ttaatttgaa ttttctgtcc taacgtttat tttgttttct tggttatgac tgatctgaaa
     4861 ttaatttttg aattttaagg taatatgtca ggcggtgccg caagtttgta caaaaaagca
     4921 ggctccatga aaaagcctga actcaccgcg acgtctgtcg agaagtttct gatcgaaaag
     4981 ttcgacagcg tctccgacct gatgcagctc tcggagggcg aagaatctcg tgctttcagc
     5041 ttcgatgtag gagggcgtgg atatgtcctg cgggtaaata gctgcgccga tggtttctac
     5101 aaagatcgtt atgtttatcg gcactttgca tcggccgcgc tcccgattcc ggaagtgctt
     5161 gacattgggg aattcagcga gagcctgacc tattgcatct cccgccgtgc acagggtgtc
     5221 acgttgcaag acctgcctga aaccgaactg cccgctgttc tgcagccggt cgcggaggcc
     5281 atggatgcga tcgctgcggc cgatcttagc cagacgagcg ggttcggccc attcggaccg
     5341 caaggaatcg gtcaatacac tacatggcgt gatttcatat gcgcgattgc tgatccccat
     5401 gtgtatcact ggcaaactgt gatggacgac accgtcagtg cgtccgtcgc gcaggctctc
     5461 gatgagctga tgctttgggc cgaggactgc cccgaagtcc ggcacctcgt gcacgcggat
     5521 ttcggctcca acaatgtcct gacggacaat ggccgcataa cagcggtcat tgactggagc
     5581 gaggcgatgt tcggggattc ccaatacgag gtcgccaaca tcttcttctg gaggccgtgg
     5641 ttggcttgta tggagcagca gacgcgctac ttcgagcgga ggcatccgga gcttgcagga
     5701 tcgccgcggc tccgggcgta tatgctccgc attggtcttg accaactcta tcagagcttg
     5761 gttgacggca atttcgatga tgcagcttgg gcgcagggtc gatgcgacgc aatcgtccga
     5821 tccggagccg ggactgtcgg gcgtacacaa atcgcccgca gaagcgcggc cgtctggacc
     5881 gatggctgtg tagaagtact cgccgatagt ggaaaccgac gccccagcac tcgtccgagg
     5941 gcaaaggaat agacccagct ttcttgtaca aagtgggtcc aattactctt caacatccct
     6001 acatgctctt tctccctgtg ctcccacccc ctatttttgt tattatcaaa aaacttctct
     6061 taatttcttt gttttttagc ttcttttaag tcacctctaa caatgaaatt gtgtagattc
     6121 aaaaatagaa ttaattcgta ataaaaagtc gaaaaaaatt gtgctccctc cccccattaa
     6181 taataattct atcccaaaat ctacacaatg ttctgtgtac acttcttatg ttttttactt
     6241 ctgataaatt tttttgaaac atcatagaaa aaaccgcaca caaaatacct tatcatatgt
     6301 tacgtttcag tttatgaccg caatttttaa taacttcgta taggatactt tatacgaagt
     6361 tattttcagg gtcgaagagc caactttatt atacaaagtt ggcattataa aaaagcattg
     6421 cttatcaatt tgttgcaacg aacaggtcac tatcagtcaa aataaaatca ttatttggag
     6481 ctccatggta gcgttaacgc ggccgcgata tcccctatag tgagtcgtat tacatggtca
     6541 tagctgtttc ctggcagctc tggcccgtgt ctcaaaatct ctgatgttac attgcacaag
     6601 ataaaaatat atcatcatga acaataaaac tgtctgctta cataaacagt aatacaaggg
     6661 gtgttatgag ccatattcaa cgggaaacgt cgaggccgcg attaaattcc aacatggatg
     6721 ctgatttata tgggtataaa tgggctcgcg ataatgtcgg gcaatcaggt gcgacaatct
     6781 atcgcttgta tgggaagccc gatgcgccag agttgtttct gaaacatggc aaaggtagcg
     6841 ttgccaatga tgttacagat gagatggtca gactaaactg gctgacggaa tttatgcctc
     6901 ttccgaccat caagcatttt atccgtactc ctgatgatgc atggttactc accactgcga
     6961 tccccggaaa aacagcattc caggtattag aagaatatcc tgattcaggt gaaaatattg
     7021 ttgatgcgct ggcagtgttc ctgcgccggt tgcattcgat tcctgtttgt aattgtcctt
     7081 ttaacagcga tcgcgtattt cgtctcgctc aggcgcaatc acgaatgaat aacggtttgg
     7141 ttgatgcgag tgattttgat gacgagcgta atggctggcc tgttgaacaa gtctggaaag
     7201 aaatgcataa acttttgcca ttctcaccgg attcagtcgt cactcatggt gatttctcac
     7261 ttgataacct tatttttgac gaggggaaat taataggttg tattgatgtt ggacgagtcg
     7321 gaatcgcaga ccgataccag gatcttgcca tcctatggaa ctgcctcggt gagttttctc
     7381 cttcattaca gaaacggctt tttcaaaaat atggtattga taatcctgat atgaataaat
     7441 tgcagtttca tttgatgctc gatgagtttt tctaatcaga attggttaat tggttgtaac
     7501 actggcagag cattacgctg acttgacggg acggcgcaag ctcatgacca aaatccctta
     7561 acgtgagtta cgcgtcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt
     7621 cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac
     7681 cagcggtggt ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct
     7741 tcagcagagc gcagatacca aatactgttc ttctagtgta gccgtagtta ggccaccact
     7801 tcaagaactc tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg
     7861 ctgccagtgg cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata
     7921 aggcgcagcg gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga
     7981 cctacaccga actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag
     8041 ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg
     8101 agcttccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac
     8161 ttgagcgtcg atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca
     8221 acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg tt
//
