LOCUS       Exported                3745 bp ds-DNA     circular SYN 28-JUN-2022
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  recombinant plasmid
REFERENCE   1  (bases 1 to 3745)
  AUTHORS   Wythe Lab
  TITLE     Direct Submission
  JOURNAL   Exported Monday, Jul 22, 2024 from SnapGene 5.0.8
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..3745
                     /organism="recombinant plasmid"
                     /mol_type="other DNA"
     terminator      268..295
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      387..473
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     misc_feature    complement(481..498)
                     /label=SeqLa
     misc_feature    537..553
                     /label=M13 For -20
     protein_bind    569..668
                     /gene="mutant version of attL"
                     /label=attL1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     645..691
                     /label=oJDW 2008 (pME 3xNLS-Cre FWD)
     misc_feature    685..691
                     /label=Kozak
     CDS             692..694
                     /codon_start=1
                     /translation="M"
     CDS             695..715
                     /codon_start=1
                     /product="nuclear localization signal of SV40 large T 
                     antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             719..739
                     /codon_start=1
                     /product="nuclear localization signal of SV40 large T 
                     antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             743..763
                     /codon_start=1
                     /product="nuclear localization signal of SV40 large T 
                     antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             764..778
                     /codon_start=1
                     /product="Gly Gly Ala Gly Gly Flexible Linker"
                     /label=G2AG2 Linker
                     /label=3X Flag GlyGlyAlaGlyGly
                     /note="Sabourin M et al., A flexible protein linker 
                     improves the function of epitope-tagged proteins in 
                     Saccharomyces cerevisiae (2007). Yeast."
                     /translation="GGAGG"
     CDS             812..826
                     /codon_start=1
                     /product="Gly Gly Ala Gly Gly Flexible Linker"
                     /label=G2AG2 Linker
                     /label=3X Flag GlyGlyAlaGlyGly
                     /note="Sabourin M et al., A flexible protein linker 
                     improves the function of epitope-tagged proteins in 
                     Saccharomyces cerevisiae (2007). Yeast."
                     /translation="GGAGG"
     CDS             827..1858
                     /codon_start=1
                     /product="site-specific recombinase"
                     /label=Cre
                     /note="Cre is a site-specific recombinase from 
                     bacteriophage P1. Recombination occurs at loxP sequences."
                     /translation="MANLLTVHQNLPALPVDATSDEVRKNLMDMFRDRQAFSEHTWKML
                     LSVCRSWAAWCKLNNRKWFPAEPEDVRDYLLYLQARGLAVKTIQQHLGQLNMLHRRSGL
                     PRPSDSNAVSLVMRRIRKENVDAGERAKQALAFERTDFDQVRSLMENSDRCQDIRNLAF
                     LGIAYNTLLRIAEIARIRVKDISRTDGGRMLIHIGRTKTLVSTAGVEKALSLGVTKLVE
                     RWISVSGVADDPNNYLFCRVRKNGVAAPSATSQLSTRALEGIFEATHRLIYGAKDDSGQ
                     RYLAWSGHSARVGAARDMARAGVSIPEIMQAGGWTNVNIVMNYIRNLDSETGAMVRLLE
                     DGD"
     primer_bind     complement(1843..1891)
                     /label=oJDW 2009 (pME 3xNLS-Cre REV)
     protein_bind    complement(1870..1969)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        complement(1987..2005)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2010..2026)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_feature    2040..2061
                     /label=SeqL B
     CDS             2139..2948
                     /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"
     rep_origin      3095..3683
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
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 tcgggcccca aataatgatt ttattttgac tgatagtgac
      601 ctgttcgttg caacacattg atgagcaatg cttttttata atgccaactt tgtacaaaaa
      661 agcaggctct cgagttattc tagacgccac catgccaaaa aagaagagaa aggtaggtcc
      721 aaaaaagaag agaaaggtag gtccaaaaaa gaagagaaag gtaggtggtg ctggtggtag
      781 atctgagctc ttctgcagtc gacgcgggcc cggtggtgct ggtggtatgg ccaatttact
      841 gaccgtacac caaaatttgc ctgcattacc ggtcgatgca acgagtgatg aggttcgcaa
      901 gaacctgatg gacatgttca gggatcgcca ggcgttttct gagcatacct ggaaaatgct
      961 tctgtccgtt tgccggtcgt gggcggcatg gtgcaagttg aataaccgga aatggtttcc
     1021 cgcagaacct gaagatgttc gcgattatct tctatatctt caggcgcgcg gtctggcagt
     1081 aaaaactatc cagcaacatt tgggccagct aaacatgctt catcgtcggt ccgggctgcc
     1141 acgaccaagt gacagcaatg ctgtttcact ggttatgcgg cggatccgaa aagaaaacgt
     1201 tgatgccggt gaacgtgcaa aacaggctct agcgttcgaa cgcactgatt tcgaccaggt
     1261 tcgttcactc atggaaaata gcgatcgctg ccaggatata cgtaatctgg catttctggg
     1321 gattgcttat aacaccctgt tacgtatagc cgaaattgcc aggatcaggg ttaaagatat
     1381 ctcacgtact gacggtggga gaatgttaat ccatattggc agaacgaaaa cgctggttag
     1441 caccgcaggt gtagagaagg cacttagcct gggggtaact aaactggtcg agcgatggat
     1501 ttccgtctct ggtgtagctg atgatccgaa taactacctg ttttgccggg tcagaaaaaa
     1561 tggtgttgcc gcgccatctg ccaccagcca gctatcaact cgcgccctgg aagggatttt
     1621 tgaagcaact catcgattga tttacggcgc taaggatgac tctggtcaga gatacctggc
     1681 ctggtctgga cacagtgccc gtgtcggagc cgcgcgagat atggcccgcg ctggagtttc
     1741 aataccggag atcatgcaag ctggtggctg gaccaatgta aatattgtca tgaactatat
     1801 ccgtaacctg gatagtgaaa caggggcaat ggtgcgcctg ctggaagatg gcgattagta
     1861 gtagactagt cccagctttc ttgtacaaag ttggcattat aagaaagcat tgcttatcaa
     1921 tttgttgcaa cgaacaggtc actatcagtc aaaataaaat cattatttgc catccagctg
     1981 atatccccta tagtgagtcg tattacatgg tcatagctgt ttcctggcag ctctggcccg
     2041 tgtctcaaaa tctctgatgt tacattgcac aagataaaat aatatcatca tgaacaataa
     2101 aactgtctgc ttacataaac agtaatacaa ggggtgttat gagccatatt caacgggaaa
     2161 cgtcgaggcc gcgattaaat tccaacatgg atgctgattt atatgggtat aaatgggctc
     2221 gcgataatgt cgggcaatca ggtgcgacaa tctatcgctt gtatgggaag cccgatgcgc
     2281 cagagttgtt tctgaaacat ggcaaaggta gcgttgccaa tgatgttaca gatgagatgg
     2341 tcagactaaa ctggctgacg gaatttatgc ctcttccgac catcaagcat tttatccgta
     2401 ctcctgatga tgcatggtta ctcaccactg cgatccccgg aaaaacagca ttccaggtat
     2461 tagaagaata tcctgattca ggtgaaaata ttgttgatgc gctggcagtg ttcctgcgcc
     2521 ggttgcattc gattcctgtt tgtaattgtc cttttaacag cgatcgcgta tttcgtctcg
     2581 ctcaggcgca atcacgaatg aataacggtt tggttgatgc gagtgatttt gatgacgagc
     2641 gtaatggctg gcctgttgaa caagtctgga aagaaatgca taaacttttg ccattctcac
     2701 cggattcagt cgtcactcat ggtgatttct cacttgataa ccttattttt gacgagggga
     2761 aattaatagg ttgtattgat gttggacgag tcggaatcgc agaccgatac caggatcttg
     2821 ccatcctatg gaactgcctc ggtgagtttt ctccttcatt acagaaacgg ctttttcaaa
     2881 aatatggtat tgataatcct gatatgaata aattgcagtt tcatttgatg ctcgatgagt
     2941 ttttctaatc agaattggtt aattggttgt aacactggca gagcattacg ctgacttgac
     3001 gggacggcgc aagctcatga ccaaaatccc ttaacgtgag ttacgcgtcg ttccactgag
     3061 cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa
     3121 tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag
     3181 agctaccaac tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg
     3241 ttcttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat
     3301 acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta
     3361 ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg
     3421 gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc
     3481 gtgagctatg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa
     3541 gcggcagggt cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc
     3601 tttatagtcc tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt
     3661 caggggggcg gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct
     3721 tttgctggcc ttttgctcac atgtt
//
