LOCUS       sequence-233447-        7055 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Targeting vector for mouse Ddx42 to insert 2A-tdiRFP670-PEST into 
            immediately upstream of stop codon..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7055)
  AUTHORS   Ochiai H, Hayashi T, Umeda M, Yoshimura M, Harada A, Shimizu Y, 
            Nakano K, Saitoh N, Liu Z, Yamamoto T, Okamura T, Ohkawa Y, Kimura 
            H, Nikaido I
  TITLE     Genome-wide kinetic properties of transcriptional bursting in mouse 
            embryonic stem cells.
  JOURNAL   Sci Adv. 2020 Jun 17;6(25):eaaz6699. doi: 10.1126/sciadv.aaz6699. 
            eCollection 2020 Jun.
  PUBMED    32596448
REFERENCE   2  (bases 1 to 7055)
  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: "Sci Adv."; 
            date: "2020-06-17"; volume: "6(25):eaaz6699. doi"; pages: " 
            10.1126/sciadv.aaz6699. eCollection 2020 Jun"
COMMENT     Sequence Label: pTV-Ddx42-tdiRFP670-PEST
FEATURES             Location/Qualifiers
     source          1..7055
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(30..458)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(140..161)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     352..371
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     585..607
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     599..616
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     600..616
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     626..645
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        626..644
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1643..1696
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     protein_bind    complement(1697..1730)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             1739..1759
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             3722..3841
                     /codon_start=1
                     /product="PEST degradation sequence from mouse ornithine 
                     decarboxylase"
                     /label=hPEST
                     /note="human codon-optimized"
                     /translation="SHGFPPEVEEQAAGTLPMSCAQESGMDRHPAACASARINV"
     protein_bind    3844..3877
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     primer_bind     complement(4867..4887)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(4867..4885)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(4906..4922)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4906..4922)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4919..4941)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4930..4946
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(4954..4960,4961..4978,4979..4984))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4999..5020
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5137..5154)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5308..5896)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5388..5407)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6067..6858,6859..6927))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     6690..6709
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6928..7032)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ctaaattgta agcgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc
       61 attttttaac caataggccg aaatcggcaa aatcccttat aaatcaaaag aatagaccga
      121 gatagggttg agtgttgttc cagtttggaa caagagtcca ctattaaaga acgtggactc
      181 caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg aaccatcacc
      241 ctaatcaagt tttttggggt cgaggtgccg taaagcacta aatcggaacc ctaaagggag
      301 cccccgattt agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa
      361 agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac
      421 cacacccgcc gcgcttaatg cgccgctaca gggcgcgtcc cattcgccat tcaggctgcg
      481 caactgttgg gaagggcgat cggtgcgggc ctcttcgcta ttacgccagc tggcgaaagg
      541 gggatgtgct gcaaggcgat taagttgggt aacgccaggg ttttcccagt cacgacgttg
      601 taaaacgacg gccagtgagc gcgcgtaata cgactcacta tagggcgaat tgggtaccgc
      661 tatcccgaaa gttccctata ccctccctaa aataaacctt aaaaaaaaac aacaacaaca
      721 acaactttag aacagtatct cttggaagag tgaggaaacc ttcctgagaa agcagctgag
      781 tctgtaagct tcatgctaga tttagagcct tatgttcact tgattttcac atacaggcat
      841 taacagtatc ctgcttgaat acacaagtgg agtttctgta cactttatca ttaacgatta
      901 tttgtaacat tatttgtata ttgaagtttg ttatacttat tcctggtgtc tttttttttt
      961 tttctttcag tcccagtaca agagtcactt tgttgcagcc agtttaagta atcagaaggc
     1021 cggaacctct tctgcagggg caagtggatg gactagtgca ggaagcttaa attctgttcc
     1081 aactaattca gcacagcagg gccataatag tcctgacaac cccatgacca gttctaccaa
     1141 gaacatccca ggcttcaaca actctgggaa tatcagtagt gccccagtga cctacccttc
     1201 tattggagcc caaggagtca acaacacagc ttcagggaat aacagccgtg aagggattgg
     1261 gggtggcaat gggaagagag agagatatac tgagaaccgg ggtggtagcc gtcatagcca
     1321 tggagatggt ggcaatcggc atggagatgg tgggcgccat ggagatggat accgttatcc
     1381 agaaagtggt agccgtcata ctgatggcca ccgacatgga gagaccaggc atggaggaag
     1441 tgccggcaga cacggagaga gccggggtgc aaatgatggc aggaatggag aaagcaggaa
     1501 agaaggtttt aatcgtgaaa acaagatgga ccccaaggtg gatagcagca ggatggacaa
     1561 agtggacagc aagacagata agacacctga tggctttgct gttccagagc cacccaaaag
     1621 aaagaaaagc cgatgggata gtgagggaag aggtagcctg ctaacttgcg gggacgtaga
     1681 ggaaaatcct ggtcccataa cttcgtatag catacattat acgaagttat caatgggccc
     1741 aaaaaagaaa agaaaagttg aattcatggc cagaaaagtg gacctgacca gttgtgaccg
     1801 agagccgata cacattcctg gctctatcca gccatgtggg tgtttgctgg cctgcgatgc
     1861 ccaggcagtc aggattactc gaattaccga aaatgcgggt gccttctttg gccgtgaaac
     1921 gccccgtgtc ggcgaactct tagccgatta tttcggagaa accgaggctc acgcacttag
     1981 gaacgcactg gcgcaatcaa gcgatcccaa acgccctgct ctcatctttg ggtggagaga
     2041 tggactgaca ggccgtactt tcgacataag ccttcaccgg catgacggaa ccagtataat
     2101 tgagtttgaa ccggccgccg ctgagcaagc agacaacccc cttcgcctga ccagacaaat
     2161 tatcgcccgc actaaggagc tgaaatcgct ggaagaaatg gccgctcgtg ttcccaggta
     2221 tcttcaggca atgctgggtt atcatagggt aatgctctac agattcgccg acgatggctc
     2281 tggtatggta ataggggagg ccaagcgatc cgacctggag tcttttctgg ggcagcattt
     2341 ccccgcatct ctagtacccc agcaggcaag gcttttgtac ctcaagaatg ccatccgcgt
     2401 ggtatcggac tcccgcggta ttagtagcag gatcgttcca gaacacgacg cctctggagc
     2461 tgcgctggat ctgtctttcg ctcacctccg atctatcagc ccttgtcatt tagagtttct
     2521 gcgaaatatg ggggtcagcg caagtatgtc cctgagcata ataatcgatg gaacactatg
     2581 gggcctaata atttgccatc attacgaacc tagagcagta ccaatggcac agcgagtggc
     2641 cgccgaaatg ttcgccgatt ttttgtctct tcattttact gccgctcacc atcagcgcgg
     2701 ccacgggact ggctcgacag gttctggcag tagtgggact gcttcatcag aagataataa
     2761 catggccatg gccagaaaag tggacctgac cagttgtgac cgagagccga tacacattcc
     2821 tggctctatc cagccatgtg ggtgtttgct ggcctgcgat gcccaggcag tcaggattac
     2881 tcgaattacc gaaaatgcgg gtgccttctt tggccgtgaa acgccccgtg tcggcgaact
     2941 cttagccgat tatttcggag aaaccgaggc tcacgcactt aggaacgcac tggcgcaatc
     3001 aagcgatccc aaacgccctg ctctcatctt tgggtggaga gatggactga caggccgtac
     3061 tttcgacata agccttcacc ggcatgacgg aaccagtata attgagtttg aaccggccgc
     3121 cgctgagcaa gcagacaacc cccttcgcct gaccagacaa attatcgccc gcactaagga
     3181 gctgaaatcg ctggaagaaa tggccgctcg tgttcccagg tatcttcagg caatgctggg
     3241 ttatcatagg gtaatgctct acagattcgc cgacgatggc tctggtatgg taatagggga
     3301 ggccaagcga tccgacctgg agtcttttct ggggcagcat ttccccgcat ctctagtacc
     3361 ccagcaggca aggcttttgt acctcaagaa tgccatccgc gtggtatcgg actcccgcgg
     3421 tattagtagc aggatcgttc cagaacacga cgcctctgga gctgcgctgg atctgtcttt
     3481 cgctcacctc cgatctatca gcccttgtca tttagagttt ctgcgaaata tgggggtcag
     3541 cgcaagtatg tccctgagca taataatcga tggaacacta tggggcctaa taatttgcca
     3601 tcattacgaa cctagagcag taccaatggc acagcgagtg gccgccgaaa tgttcgccga
     3661 ttttttgtct cttcatttta ctgccgctca ccatcagcgc ggatccgcgt ttaaactcga
     3721 gtctcacggc ttccctcccg aggtggagga gcaggccgcc ggcaccctgc ccatgagctg
     3781 cgcccaggag agcggcatgg atagacaccc tgctgcttgc gccagcgcca ggatcaacgt
     3841 cttgaagttc ctattctcta gaaagtatag gaacttctag tagagggtat gtgaaaatca
     3901 gcgtcgtcag tcgtctttaa ttttattttt agaaagattt tggtaatgag gcgtctcagg
     3961 gctgggttgg ggtccaagat atacagaccc cctgcccttg gtggagaact ggagagtgga
     4021 ttgtaccttc atctggtgtt cattcagagg tttccttggg aacctgagct ggccctggaa
     4081 agcagagagc tgggaagctt ttgtttgttg tgggtgagtt ttaaggaggg taagtagaat
     4141 acttagggag aaagggcctt ttagggcaca aaaactcact ctaggtttat aatagcttat
     4201 attttttact aaaatgtcac cttataaaca tctataaatt ggactttttt tcttagctgt
     4261 ggccaggcag tcccaacttc tgccctgtta gggatgcttg gggagcagtg gtgctttcag
     4321 atgtttgcag ggaagatgcc tgagcgctca ggttccccag actgtttcca gccctgagac
     4381 acgtaggaaa gacagccttc cagacacagg ctttgaaatc ctaagccatt aggactgaag
     4441 gaatgatcat cttggaagga ttccaggata atccaggcct ggagtactgc taaatttacc
     4501 atttaaaacc aaccccaaca gtagcattca aagtgtctat ttgttttgtt ttgttttttt
     4561 aaacttgccc taatagtaga aaaaagtctt ctgctgaaat gattttgatt tttgttctat
     4621 cttgtttata aaagaaaaat atacaaactt tgaattttgt gatttttgtg aaggtttctt
     4681 taagatgtgc attccctctc tgcttgctcg atagtaaatg tcttactact gggacctgag
     4741 ggctgtcgtt tctaccatac aagttagttg ctacatatta aattatggct tggaacagta
     4801 cagcccatga tgtgaaatgt ggggaagtgc ttgcttatta aactatcaga gctccagctt
     4861 ttgttccctt tagtgagggt taattgcgcg cttggcgtaa tcatggtcat agctgtttcc
     4921 tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa gcataaagtg
     4981 taaagcctgg ggtgcctaat gagtgagcta actcacatta attgcgttgc gctcactgcc
     5041 cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg
     5101 gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc
     5161 ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac
     5221 agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa
     5281 ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca
     5341 caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc
     5401 gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata
     5461 cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta
     5521 tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca
     5581 gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga
     5641 cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg
     5701 tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagga cagtatttgg
     5761 tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg
     5821 caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag
     5881 aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa
     5941 cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat
     6001 ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc
     6061 tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc
     6121 atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc
     6181 tggccccagt gctgcaatga taccgcggct gccacgctca ccggctccag atttatcagc
     6241 aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc
     6301 catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt
     6361 gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc
     6421 ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa
     6481 aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt
     6541 atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg
     6601 cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc
     6661 gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa
     6721 agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt
     6781 gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt
     6841 caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag
     6901 ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta
     6961 tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat
     7021 aggggttccg cgcacatttc cccgaaaagt gccac
//
