LOCUS       sequence-233452-        7267 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Targeting vector for mouse Dhx15 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 7267)
  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 7267)
  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-Dhx15-tdiRFP670-PEST
FEATURES             Location/Qualifiers
     source          1..7267
                     /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             1734..1787
                     /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(1788..1821)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             1830..1850
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             3813..3932
                     /codon_start=1
                     /product="PEST degradation sequence from mouse ornithine 
                     decarboxylase"
                     /label=hPEST
                     /note="human codon-optimized"
                     /translation="SHGFPPEVEEQAAGTLPMSCAQESGMDRHPAACASARINV"
     protein_bind    3935..3968
                     /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(5079..5099)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(5079..5097)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(5118..5134)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5118..5134)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5131..5153)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5142..5158
                     /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(5166..5172,5173..5190,5191..5196))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5211..5232
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5349..5366)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5520..6108)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5600..5619)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6279..7070,7071..7139))
                     /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     6902..6921
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7140..7244)
                     /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 tgggtaccgg
      661 gccccccctc gagtagcttg aacttctcag ctaggtgtgt atattcttgt acgtggacct
      721 aggattaaca tggtatgatt tgtttttttc ttttgtttct taagtcagtg ttctggtgag
      781 aggacagtgg gaatattagt gtgggcaggg agttgagtac catcaggaat ttcttggtgc
      841 cattttgtgg tctctgttgg cagatgtttg tctttttaat attttacttg gtttaagtat
      901 tttgtcttta tttattaaat gttgaggggt tttttgacct atgtaaagat aaaaattata
      961 tcaaattatt ccctagcata ttagcttcta aaaattgtaa ttaagacagg ttgttcaatt
     1021 tttagcaaaa tggtaccctt gtgatttccc taccaccacc atttttataa gttatgggaa
     1081 tgtcagccaa gagtggacag gaaaccattt atgttggttt gtctcactta ctattttgcg
     1141 gcttccatcc attactgatt tatggctttt gacacttcaa ttgtttagaa cattaccctt
     1201 ttattatgtt ttgccaaggg aaaacagtat agtttataag atatacagtt ttatatttgt
     1261 agttcttaat actgataatg agatggagtt atttgactat caaaaccata acctttaaaa
     1321 tacaggttta agtgtaggca aatgaaacac taaatagctt ttgtcttttc atattaatgt
     1381 aaataaagta ttagaaaaat taatcccgat cctgtttcca aggtggtaag tttacagaag
     1441 gtttatactt ttgctaagct gatatttctt aaactgtgtt aataaagata tgattttaaa
     1501 tcttttatca ttgatttgta tttgaacatt tgtcgtcatt cagcagcagt ataattatta
     1561 gcaactctag tacttaaatc ttgatttcat aactttcccc cccccccatt ctggacaggt
     1621 tggtgaaaat tgcccctcaa tattatgaca tgagcaattt cccacagtgt gaagcaaaga
     1681 gacagttgga ccgcattatt gccaaacttc aatccaagga atattcacag tacgagggaa
     1741 gaggtagcct gctaacttgc ggggacgtag aggaaaatcc tggtcccata acttcgtata
     1801 gcatacatta tacgaagtta tcaatgggcc caaaaaagaa aagaaaagtt gaattcatgg
     1861 ccagaaaagt ggacctgacc agttgtgacc gagagccgat acacattcct ggctctatcc
     1921 agccatgtgg gtgtttgctg gcctgcgatg cccaggcagt caggattact cgaattaccg
     1981 aaaatgcggg tgccttcttt ggccgtgaaa cgccccgtgt cggcgaactc ttagccgatt
     2041 atttcggaga aaccgaggct cacgcactta ggaacgcact ggcgcaatca agcgatccca
     2101 aacgccctgc tctcatcttt gggtggagag atggactgac aggccgtact ttcgacataa
     2161 gccttcaccg gcatgacgga accagtataa ttgagtttga accggccgcc gctgagcaag
     2221 cagacaaccc ccttcgcctg accagacaaa ttatcgcccg cactaaggag ctgaaatcgc
     2281 tggaagaaat ggccgctcgt gttcccaggt atcttcaggc aatgctgggt tatcataggg
     2341 taatgctcta cagattcgcc gacgatggct ctggtatggt aataggggag gccaagcgat
     2401 ccgacctgga gtcttttctg gggcagcatt tccccgcatc tctagtaccc cagcaggcaa
     2461 ggcttttgta cctcaagaat gccatccgcg tggtatcgga ctcccgcggt attagtagca
     2521 ggatcgttcc agaacacgac gcctctggag ctgcgctgga tctgtctttc gctcacctcc
     2581 gatctatcag cccttgtcat ttagagtttc tgcgaaatat gggggtcagc gcaagtatgt
     2641 ccctgagcat aataatcgat ggaacactat ggggcctaat aatttgccat cattacgaac
     2701 ctagagcagt accaatggca cagcgagtgg ccgccgaaat gttcgccgat tttttgtctc
     2761 ttcattttac tgccgctcac catcagcgcg gccacgggac tggctcgaca ggttctggca
     2821 gtagtgggac tgcttcatca gaagataata acatggccat ggccagaaaa gtggacctga
     2881 ccagttgtga ccgagagccg atacacattc ctggctctat ccagccatgt gggtgtttgc
     2941 tggcctgcga tgcccaggca gtcaggatta ctcgaattac cgaaaatgcg ggtgccttct
     3001 ttggccgtga aacgccccgt gtcggcgaac tcttagccga ttatttcgga gaaaccgagg
     3061 ctcacgcact taggaacgca ctggcgcaat caagcgatcc caaacgccct gctctcatct
     3121 ttgggtggag agatggactg acaggccgta ctttcgacat aagccttcac cggcatgacg
     3181 gaaccagtat aattgagttt gaaccggccg ccgctgagca agcagacaac ccccttcgcc
     3241 tgaccagaca aattatcgcc cgcactaagg agctgaaatc gctggaagaa atggccgctc
     3301 gtgttcccag gtatcttcag gcaatgctgg gttatcatag ggtaatgctc tacagattcg
     3361 ccgacgatgg ctctggtatg gtaatagggg aggccaagcg atccgacctg gagtcttttc
     3421 tggggcagca tttccccgca tctctagtac cccagcaggc aaggcttttg tacctcaaga
     3481 atgccatccg cgtggtatcg gactcccgcg gtattagtag caggatcgtt ccagaacacg
     3541 acgcctctgg agctgcgctg gatctgtctt tcgctcacct ccgatctatc agcccttgtc
     3601 atttagagtt tctgcgaaat atgggggtca gcgcaagtat gtccctgagc ataataatcg
     3661 atggaacact atggggccta ataatttgcc atcattacga acctagagca gtaccaatgg
     3721 cacagcgagt ggccgccgaa atgttcgccg attttttgtc tcttcatttt actgccgctc
     3781 accatcagcg cggatccgcg tttaaactcg agtctcacgg cttccctccc gaggtggagg
     3841 agcaggccgc cggcaccctg cccatgagct gcgcccagga gagcggcatg gatagacacc
     3901 ctgctgcttg cgccagcgcc aggatcaacg tcttgaagtt cctattctct agaaagtata
     3961 ggaacttcta gtgaattcaa tgcttagaac tgaagttatt cagaggacag ctttaaaaga
     4021 tgaatgaagt caagttcaag ttgtgctctt catgttggtt caataatggc ctttatttga
     4081 aagcttttta atttttcttt acagtaaata ttccattctg atttcataaa ttaaacattt
     4141 atgcctccct tttgtgttga cactgtagct catactggaa aagtcgatca atgttttgca
     4201 gtttattgaa agtagttcta tatataacaa tgttataagc atttctttag aaatggttga
     4261 aaatgcttct aaaatgtgat tatcgaccat ggtatgcatg atcgttgtaa ttgttgacat
     4321 tccttttaga agttgtgaaa tgttacaact tgtgcttctg tagacacaat cttctgtctc
     4381 catacagagg cactgacttc aataaagtct atttatacta atcttggccc aagtccttct
     4441 gtatctttta ttctaagagt ctctatagtg agtggctgct tttctttttt cattgtaatt
     4501 atactgttac tacgtattta gcgtttttta aaaggagtac tccttaggta cagtagccat
     4561 taaacctggg ttttaaagag cataaatgta gcactttctt aggttaatca gtatgtaatc
     4621 taatgggata acgtgtgtaa tatttttttt tgaaattgga tccccagaga cagaataatg
     4681 gaagaatggc tttgtcctca ttcttaaagc tcatttcata aggctgatag tttattacct
     4741 tattaaatgc gttgtgtcag gtcttactgt actgaggcga acagtttaat cactctagcc
     4801 cctagcatgt gtttgtattg gataggaggg aacagtgttc tggatcaccg tcagttttta
     4861 ctaagtattg actgttggca gcttttataa tgtaatttgt atagtgtgtt aaaatgagaa
     4921 gttgagcttc ctatcaacag gcaggagaca aactaaaatc cttccggaac caaaaggagc
     4981 tgttgatgtg gccagtcaaa agtactgcaa taatttgtct ttcacttaca ggctaaatag
     5041 ttgcggccgc caccgcggtg gagctccagc ttttgttccc tttagtgagg gttaattgcg
     5101 cgcttggcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc gctcacaatt
     5161 ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta atgagtgagc
     5221 taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc
     5281 cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct
     5341 tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca
     5401 gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac
     5461 atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt
     5521 ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg
     5581 cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc
     5641 tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc
     5701 gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc
     5761 aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac
     5821 tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt
     5881 aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct
     5941 aactacggct acactagaag gacagtattt ggtatctgcg ctctgctgaa gccagttacc
     6001 ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt
     6061 ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg
     6121 atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc
     6181 atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa
     6241 tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag
     6301 gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg
     6361 tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcgg
     6421 ctgccacgct caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag
     6481 cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa
     6541 gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc
     6601 atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca
     6661 aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg
     6721 atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat
     6781 aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc
     6841 aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg
     6901 gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg
     6961 gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt
     7021 gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca
     7081 ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata
     7141 ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac
     7201 atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa
     7261 gtgccac
//
