LOCUS       sequence-227888-        4819 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Destination Vector.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4819)
  AUTHORS   Wagner TE, Becraft JR, Bodner K, Teague B, Zhang X, Woo A, Porter E,
            Alburquerque B, Dobosh B, Andries O, Sanders NN, Beal J, Densmore D,
            Kitada T, Weiss R
  TITLE     Small-molecule-based regulation of RNA-delivered circuits in 
            mammalian cells.
  JOURNAL   Nat Chem Biol. 2018 Nov;14(11):1043-1050. doi: 
            10.1038/s41589-018-0146-9. Epub 2018 Oct 16.
  PUBMED    30327560
REFERENCE   2  (bases 1 to 4819)
  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"; doi: 
            "10.1038/s41589-018-0146-9"; journalName: "Nat Chem Biol"; date: 
            "2018-11"; volume: "14"; issue: "11"; pages: "1043-1050"
COMMENT     Sequence Label: pTW325
FEATURES             Location/Qualifiers
     source          1..4819
                     /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"
     primer_bind     572..592
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    606..837
                     /gene="mutant version of attP"
                     /label=attP1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             complement(1233..1538)
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     complement(1335..1354)
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             complement(1858..2541)
                     /codon_start=1
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGRKRR
                     VDPAY"
     primer_bind     2456..2475
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(2542..2644)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     protein_bind    complement(2789..3020)
                     /gene="mutant version of attP"
                     /label=attP2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     complement(3060..3079)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3061..3079)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3084..3100)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3084..3100)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3202..3221)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             3213..4022
                     /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(3280..3299)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      4169..4757
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4658..4677
                     /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 tcggctcttc acgcaaatgg gcggtaggcg tggactggag
      601 accccaaata atgattttat tttgactgat agtgacctgt tcgttgcaac aaattgatga
      661 gcaatgcttt tttataatgc caactttgta caaaaaagct gaacgagaaa cgtaaaatga
      721 tataaatatc aatatattaa attagatttt gcataaaaaa cagactacat aatactgtaa
      781 aacacaacat atccagtcac tatgaatcaa ctacttagat ggtattagtg acctgtagtc
      841 gaccgacagc cttccaaatg ttcttcgggt gatgctgcca acttagtcga ccgacagcct
      901 tccaaatgtt cttctcaaac ggaatcgtcg tatccagcct actcgctatt gtcctcaatg
      961 ccgtattaaa tcataaaaag aaataagaaa aagaggtgcg agcctctttt ttgtgtgaca
     1021 aaataaaaac atctacctat tcatatacgc tagtgtcata gtcctgaaaa tcatctgcat
     1081 caagaacaat ttcacaactc ttatactttt ctcttacaag tcgttcggct tcatctggat
     1141 tttcagcctc tatacttact aaacgtgata aagtttctgt aatttctact gtatcgacct
     1201 gcagactggc tgtgtataag ggagcctgac atttatattc cccagaacat caggttaatg
     1261 gcgtttttga tgtcattttc gcggtggctg agatcagcca cttcttcccc gataacggag
     1321 accggcacac tggccatatc ggtggtcatc atgcgccagc tttcatcccc gatatgcacc
     1381 accgggtaaa gttcacggga gactttatct gacagcagac gtgcactggc cagggggatc
     1441 accatccgtc gcccgggcgt gtcaataata tcactctgta catccacaaa cagacgataa
     1501 cggctctctc ttttataggt gtaaacctta aactgcattt caccagcccc tgttctcgtc
     1561 agcaaaagag ccgttcattt caataaaccg ggcgacctca gccatccctt cctgattttc
     1621 cgctttccag cgttcggcac gcagacgacg ggcttcattc tgcatggttg tgcttaccag
     1681 accggagata ttgacatcat atatgccttg agcaactgat agctgtcgct gtcaactgtc
     1741 actgtaatac gctgcttcat agcatacctc tttttgacat acttcgggta tacatatcag
     1801 tatatattct tataccgcaa aaatcagcgc gcaaatacgc atactgttat ctggctttta
     1861 gtaagccgga tccacgcggc gtttacgccc cccctgccac tcatcgcagt actgttgtaa
     1921 ttcattaagc attctgccga catggaagcc atcacaaacg gcatgatgaa cctgaatcgc
     1981 cagcggcatc agcaccttgt cgccttgcgt ataatatttg cccatggtga aaacgggggc
     2041 gaagaagttg tccatattgg ccacgtttaa atcaaaactg gtgaaactca cccagggatt
     2101 ggctgagacg aaaaacatat tctcaataaa ccctttaggg aaataggcca ggttttcacc
     2161 gtaacacgcc acatcttgcg aatatatgtg tagaaactgc cggaaatcgt cgtggtattc
     2221 actccagagc gatgaaaacg tttcagtttg ctcatggaaa acggtgtaac aagggtgaac
     2281 actatcccat atcaccagct caccgtcttt cattgccata cggaattccg gatgagcatt
     2341 catcaggcgg gcaagaatgt gaataaaggc cggataaaac ttgtgcttat ttttctttac
     2401 ggtctttaaa aaggccgtaa tatccagctg aacggtctgg ttataggtac attgagcaac
     2461 tgactgaaat gcctcaaaat gttctttacg atgccattgg gatatatcaa cggtggtata
     2521 tccagtgatt tttttctcca ttttagcttc cttagctcct gaaaatctcg ataactcaaa
     2581 aaatacgccc ggtagtgatc ttatttcatt atggtgaaag ttggaacctc ttacgtgccg
     2641 atcaacgtct cattttcgcc aaaagttggc ccagggcttc ccggtatcaa cagggacacc
     2701 aggatttatt tattctgcga agtgatcttc cgtcacaggt atttattcgg cgcaaagtgc
     2761 gtcgggtgat gctgccaact tagtcgacta caggtcacta ataccatcta agtagttgat
     2821 tcatagtgac tggatatgtt gtgttttaca gtattatgta gtctgttttt tatgcaaaat
     2881 ctaatttaat atattgatat ttatatcatt ttacgtttct cgttcagctt tcttgtacaa
     2941 agttggcatt ataagaaagc attgcttatc aatttgttgc aacgaacagg tcactatcag
     3001 tcaaaataaa atcattattt gccatccagc tgatggtctc cttcataggg aagagcatcc
     3061 cctatagtga gtcgtattac atggtcatag ctgtttcctg gcagctctgg cccgtgtctc
     3121 aaaatctctg atgttacatt gcacaagata aaaatatatc atcatgaaca ataaaactgt
     3181 ctgcttacat aaacagtaat acaaggggtg ttatgagcca tattcaacgg gaaacgtcga
     3241 ggccgcgatt aaattccaac atggatgctg atttatatgg gtataaatgg gctcgcgata
     3301 atgtcgggca atcaggtgcg acaatctatc gcttgtatgg gaagcccgat gcgccagagt
     3361 tgtttctgaa acatggcaaa ggtagcgttg ccaatgatgt tacagatgag atggtcagac
     3421 taaactggct gacggaattt atgcctcttc cgaccatcaa gcattttatc cgtactcctg
     3481 atgatgcatg gttactcacc actgcgatcc ccggaaaaac agcattccag gtattagaag
     3541 aatatcctga ttcaggtgaa aatattgttg atgcgctggc agtgttcctg cgccggttgc
     3601 attcgattcc tgtttgtaat tgtcctttta acagcgatcg cgtatttcgt ctcgctcagg
     3661 cgcaatcacg aatgaataac ggtttggttg atgcgagtga ttttgatgac gagcgtaatg
     3721 gctggcctgt tgaacaagtc tggaaagaaa tgcataaact tttgccattc tcaccggatt
     3781 cagtcgtcac tcatggtgat ttctcacttg ataaccttat ttttgacgag gggaaattaa
     3841 taggttgtat tgatgttgga cgagtcggaa tcgcagaccg ataccaggat cttgccatcc
     3901 tatggaactg cctcggtgag ttttctcctt cattacagaa acggcttttt caaaaatatg
     3961 gtattgataa tcctgatatg aataaattgc agtttcattt gatgctcgat gagtttttct
     4021 aatcagaatt ggttaattgg ttgtaacact ggcagagcat tacgctgact tgacgggacg
     4081 gcgcaagctc atgaccaaaa tcccttaacg tgagttacgc gtcgttccac tgagcgtcag
     4141 accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct
     4201 gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac
     4261 caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat actgttcttc
     4321 tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg
     4381 ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt
     4441 tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt
     4501 gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc
     4561 tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca
     4621 gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata
     4681 gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg
     4741 ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct
     4801 ggccttttgc tcacatgtt
//
