LOCUS       sequence-233457-        7086 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Targeting vector for mouse Jade1 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 7086)
  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 7086)
  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-Jade1-tdiRFP670-PEST
FEATURES             Location/Qualifiers
     source          1..7086
                     /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             1672..1725
                     /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(1726..1759)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             1768..1788
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             3751..3870
                     /codon_start=1
                     /product="PEST degradation sequence from mouse ornithine 
                     decarboxylase"
                     /label=hPEST
                     /note="human codon-optimized"
                     /translation="SHGFPPEVEEQAAGTLPMSCAQESGMDRHPAACASARINV"
     protein_bind    3873..3906
                     /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(4898..4918)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(4898..4916)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(4937..4953)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4937..4953)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4950..4972)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4961..4977
                     /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(4985..4991,4992..5009,5010..5015))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5030..5051
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5168..5185)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5339..5927)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5419..5438)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6098..6889,6890..6958))
                     /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     6721..6740
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6959..7063)
                     /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 gagtaaaaca aagtgctgct gctgcttgct tgggtgggaa gtttgtataa
      721 tgatgtttcc agtctagaat gttatgagtt tgggtttgct tatgtaattt ttcctctggt
      781 tttctgttcc taggtgtgcc ttcttcctgc tcctccgcac tggagaacat gctttttttc
      841 aacagtcctt ctgtgggccc caatgctccc aagatagagg acttgaaatg gcattctgca
      901 ttcttcagga aacaaatggg cacttccttg gttcatccac tgaaaaagtc ccataaacga
      961 gatgcagtgc agaatagttc tgggaccgag ggcaagacct cgcataagca gccaggtctt
     1021 tgtggtagaa gggaggggct ggaggtctca gaaagcttgc tgagcttaga aaagactttt
     1081 gcagaagcac gtctcctatc atcagcacaa cagaaaaatg gtgtggtgac cccagaccat
     1141 gggaaaagaa gagacaatcg ttttcattgt gatctcgtta aaggagactt aaaggacaaa
     1201 tcttttaagc agagtcacaa gcctctcagg tccacagaca catcccagag gcatctggac
     1261 aacacaagag ctgccacctc ccctggagta gggcagtcag cacctggcac caggaaggag
     1321 attgtgccca agtgcaatgg ctccctagtc aaagtgccta taacacctgc cagcccagtg
     1381 aaaagctggg gaggattccg gattccaaag aagggggagc ggcagcagca aggagaagcc
     1441 catgatgggg cctgccacca gcactcagac tgctcccatc tgggtgtaag ccgagctcca
     1501 gccaaggaga gagcgaagag caggttaaga gctgacagtg agaatgatgg gtacgcccct
     1561 gatggggaaa tgagtgactc agagagcgag gcatcagaga agaaatgtat ccatgccagc
     1621 agcaccatca gcaggaggac agatatcatc aggcgaagca tcttggcctc tgagggaaga
     1681 ggtagcctgc taacttgcgg ggacgtagag gaaaatcctg gtcccataac ttcgtatagc
     1741 atacattata cgaagttatc aatgggccca aaaaagaaaa gaaaagttga attcatggcc
     1801 agaaaagtgg acctgaccag ttgtgaccga gagccgatac acattcctgg ctctatccag
     1861 ccatgtgggt gtttgctggc ctgcgatgcc caggcagtca ggattactcg aattaccgaa
     1921 aatgcgggtg ccttctttgg ccgtgaaacg ccccgtgtcg gcgaactctt agccgattat
     1981 ttcggagaaa ccgaggctca cgcacttagg aacgcactgg cgcaatcaag cgatcccaaa
     2041 cgccctgctc tcatctttgg gtggagagat ggactgacag gccgtacttt cgacataagc
     2101 cttcaccggc atgacggaac cagtataatt gagtttgaac cggccgccgc tgagcaagca
     2161 gacaaccccc ttcgcctgac cagacaaatt atcgcccgca ctaaggagct gaaatcgctg
     2221 gaagaaatgg ccgctcgtgt tcccaggtat cttcaggcaa tgctgggtta tcatagggta
     2281 atgctctaca gattcgccga cgatggctct ggtatggtaa taggggaggc caagcgatcc
     2341 gacctggagt cttttctggg gcagcatttc cccgcatctc tagtacccca gcaggcaagg
     2401 cttttgtacc tcaagaatgc catccgcgtg gtatcggact cccgcggtat tagtagcagg
     2461 atcgttccag aacacgacgc ctctggagct gcgctggatc tgtctttcgc tcacctccga
     2521 tctatcagcc cttgtcattt agagtttctg cgaaatatgg gggtcagcgc aagtatgtcc
     2581 ctgagcataa taatcgatgg aacactatgg ggcctaataa tttgccatca ttacgaacct
     2641 agagcagtac caatggcaca gcgagtggcc gccgaaatgt tcgccgattt tttgtctctt
     2701 cattttactg ccgctcacca tcagcgcggc cacgggactg gctcgacagg ttctggcagt
     2761 agtgggactg cttcatcaga agataataac atggccatgg ccagaaaagt ggacctgacc
     2821 agttgtgacc gagagccgat acacattcct ggctctatcc agccatgtgg gtgtttgctg
     2881 gcctgcgatg cccaggcagt caggattact cgaattaccg aaaatgcggg tgccttcttt
     2941 ggccgtgaaa cgccccgtgt cggcgaactc ttagccgatt atttcggaga aaccgaggct
     3001 cacgcactta ggaacgcact ggcgcaatca agcgatccca aacgccctgc tctcatcttt
     3061 gggtggagag atggactgac aggccgtact ttcgacataa gccttcaccg gcatgacgga
     3121 accagtataa ttgagtttga accggccgcc gctgagcaag cagacaaccc ccttcgcctg
     3181 accagacaaa ttatcgcccg cactaaggag ctgaaatcgc tggaagaaat ggccgctcgt
     3241 gttcccaggt atcttcaggc aatgctgggt tatcataggg taatgctcta cagattcgcc
     3301 gacgatggct ctggtatggt aataggggag gccaagcgat ccgacctgga gtcttttctg
     3361 gggcagcatt tccccgcatc tctagtaccc cagcaggcaa ggcttttgta cctcaagaat
     3421 gccatccgcg tggtatcgga ctcccgcggt attagtagca ggatcgttcc agaacacgac
     3481 gcctctggag ctgcgctgga tctgtctttc gctcacctcc gatctatcag cccttgtcat
     3541 ttagagtttc tgcgaaatat gggggtcagc gcaagtatgt ccctgagcat aataatcgat
     3601 ggaacactat ggggcctaat aatttgccat cattacgaac ctagagcagt accaatggca
     3661 cagcgagtgg ccgccgaaat gttcgccgat tttttgtctc ttcattttac tgccgctcac
     3721 catcagcgcg gatccgcgtt taaactcgag tctcacggct tccctcccga ggtggaggag
     3781 caggccgccg gcaccctgcc catgagctgc gcccaggaga gcggcatgga tagacaccct
     3841 gctgcttgcg ccagcgccag gatcaacgtc ttgaagttcc tattctctag aaagtatagg
     3901 aacttctagt gatgaaacag ccggtgtgga agcacatggc ttgttgccgt gtgctgagga
     3961 gagctctgat aataggggca agcaaatacc cattgtttct gagctacaga catacttcct
     4021 tgcaattcag attgaatttt tttccagagt catttataaa tcattgtgag aagtttgtgt
     4081 tatttgcaac ttgttgagga aacagaagag tagactgtaa ccataagaca ctaagattag
     4141 aacctgaact aaacactata aaaaaaaaac caacaaatga agtaactttg cagagggcca
     4201 ggcttgaata gcaccatgta ttattgccct ctacattctc cccaaagcac aaactcttga
     4261 atactcactc ttacagaatc atagaattct gtgtggtgaa caagtcctat ttctagaagc
     4321 cctcatgtac catttatagc ccacaaagct tattttctag gaatggtaaa tgctgacggc
     4381 atttggctcc caggctgtgt ggtatgagga gctgaggttc tgaagcttca caacactggg
     4441 atcttgcaca tgtggcatcc aggggaagag tttacacaac aggcattgct ggaaggctgc
     4501 agggacagag cagagctgct gcactgagat ttggggcccg tgtagcaagg tcccgaagcc
     4561 ctgagtgggc tgtggacaca ggacgggtgg aggaaaagtt gtccagaaac ttctgtgaga
     4621 cagggcagct gaggcccagg tgttactagt ccgcttgacc ttcctctttc ctaactgctg
     4681 gcctttggat ccaaatggga tttccctggc tactgccagt acttggccat ctggcttcca
     4741 ataatacaat ggttattcaa gttcaagtga tgaacttctg gtgactctca ctctacagag
     4801 tcaaccacta gtgcatattt gggaaatcca ggcttccacc atgagtggat tccttaagca
     4861 agcggccgcc accgcggtgg agctccagct tttgttccct ttagtgaggg ttaattgcgc
     4921 gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc
     4981 cacacaacat acgagccgga agcataaagt gtaaagcctg gggtgcctaa tgagtgagct
     5041 aactcacatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc
     5101 agctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt
     5161 ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag
     5221 ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca
     5281 tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt
     5341 tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc
     5401 gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct
     5461 ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg
     5521 tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca
     5581 agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact
     5641 atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta
     5701 acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta
     5761 actacggcta cactagaagg acagtatttg gtatctgcgc tctgctgaag ccagttacct
     5821 tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt
     5881 tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga
     5941 tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca
     6001 tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat
     6061 caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg
     6121 cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt
     6181 agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcggc
     6241 tgccacgctc accggctcca gatttatcag caataaacca gccagccgga agggccgagc
     6301 gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag
     6361 ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca
     6421 tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa
     6481 ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga
     6541 tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata
     6601 attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca
     6661 agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg
     6721 ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg
     6781 ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg
     6841 cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag
     6901 gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac
     6961 tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca
     7021 tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag
     7081 tgccac
//
