LOCUS       sequence-233466-        7129 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Targeting vector for mouse Hdac1 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 7129)
  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 7129)
  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-mHdac1-tdiRFP670-PEST2
FEATURES             Location/Qualifiers
     source          1..7129
                     /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             1600..1653
                     /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(1654..1687)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             1696..1716
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             3679..3798
                     /codon_start=1
                     /product="PEST degradation sequence from mouse ornithine 
                     decarboxylase"
                     /label=hPEST
                     /note="human codon-optimized"
                     /translation="SHGFPPEVEEQAAGTLPMSCAQESGMDRHPAACASARINV"
     protein_bind    3801..3834
                     /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(4941..4961)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(4941..4959)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(4980..4996)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4980..4996)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4993..5015)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5004..5020
                     /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(5028..5034,5035..5052,5053..5058))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5073..5094
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5211..5228)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5382..5970)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5462..5481)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6141..6932,6933..7001))
                     /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     6764..6783
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7002..7106)
                     /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 tgggtacccc
      661 ccttgctctc ataccaatgt ttctctctct gctgcctccc agtctgctcc tctgacaaac
      721 gcattgcctg tgaggaagag ttctcggact cagatgagga gggagaaggt ggtcgcaaga
      781 actcttctaa cttcaaaaaa gccaaaagag ttaaaacaga ggatgagaaa gagaaagatc
      841 ctgaagagaa aaaaggtgag tttggattag atcaggaagg agctgaccca gacccagcca
      901 gctcctttgg agcttaaact ggaggtttga gtcttgagtc tttcagtggc tcctagaggt
      961 ggtgagaacc acagaacagt gacagtgaca ctcttggtct ctgctggctg actgaccctg
     1021 gctcacatac cacccaagcc cttttcccag tttcccaggg ggctgcctct gcctgtgttc
     1081 ttgctgaaac aggctgggaa accggtctga cctgttttca gccaacctca ccagaggacc
     1141 tgggataggc ttttctcatt tttctataac cagtgaactg tggtattgaa gaaggggttt
     1201 caggatagag gagtccgact acctgagaag ccttcaagca tacctgttct cttattttct
     1261 agaagtcaca gaagaagaga aaaccaagga ggagaagcca gaagccaaag ggtgaggaaa
     1321 ggggctggtg gctctctgta gcactagagc atcatcacct tctgtgtcct acctctgtga
     1381 gagggaagga agagcaggga cagctgctgg tccagcctac agagagaggc ctctttaagg
     1441 gtggagtctg tatgtctctg cccagttccc agcagtcacc tgggcagcca gggtctatgg
     1501 aagtcactat agccatgttt gtgcttcagg gccactagca tgagttttct catgcctgtc
     1561 tgttctctct gcagggtcaa tgaggaagtt aaactggccg agggaagagg tagcctgcta
     1621 acttgcgggg acgtagagga aaatcctggt cccataactt cgtatagcat acattatacg
     1681 aagttatcaa tgggcccaaa aaagaaaaga aaagttgaat tcatggccag aaaagtggac
     1741 ctgaccagtt gtgaccgaga gccgatacac attcctggct ctatccagcc atgtgggtgt
     1801 ttgctggcct gcgatgccca ggcagtcagg attactcgaa ttaccgaaaa tgcgggtgcc
     1861 ttctttggcc gtgaaacgcc ccgtgtcggc gaactcttag ccgattattt cggagaaacc
     1921 gaggctcacg cacttaggaa cgcactggcg caatcaagcg atcccaaacg ccctgctctc
     1981 atctttgggt ggagagatgg actgacaggc cgtactttcg acataagcct tcaccggcat
     2041 gacggaacca gtataattga gtttgaaccg gccgccgctg agcaagcaga caaccccctt
     2101 cgcctgacca gacaaattat cgcccgcact aaggagctga aatcgctgga agaaatggcc
     2161 gctcgtgttc ccaggtatct tcaggcaatg ctgggttatc atagggtaat gctctacaga
     2221 ttcgccgacg atggctctgg tatggtaata ggggaggcca agcgatccga cctggagtct
     2281 tttctggggc agcatttccc cgcatctcta gtaccccagc aggcaaggct tttgtacctc
     2341 aagaatgcca tccgcgtggt atcggactcc cgcggtatta gtagcaggat cgttccagaa
     2401 cacgacgcct ctggagctgc gctggatctg tctttcgctc acctccgatc tatcagccct
     2461 tgtcatttag agtttctgcg aaatatgggg gtcagcgcaa gtatgtccct gagcataata
     2521 atcgatggaa cactatgggg cctaataatt tgccatcatt acgaacctag agcagtacca
     2581 atggcacagc gagtggccgc cgaaatgttc gccgattttt tgtctcttca ttttactgcc
     2641 gctcaccatc agcgcggcca cgggactggc tcgacaggtt ctggcagtag tgggactgct
     2701 tcatcagaag ataataacat ggccatggcc agaaaagtgg acctgaccag ttgtgaccga
     2761 gagccgatac acattcctgg ctctatccag ccatgtgggt gtttgctggc ctgcgatgcc
     2821 caggcagtca ggattactcg aattaccgaa aatgcgggtg ccttctttgg ccgtgaaacg
     2881 ccccgtgtcg gcgaactctt agccgattat ttcggagaaa ccgaggctca cgcacttagg
     2941 aacgcactgg cgcaatcaag cgatcccaaa cgccctgctc tcatctttgg gtggagagat
     3001 ggactgacag gccgtacttt cgacataagc cttcaccggc atgacggaac cagtataatt
     3061 gagtttgaac cggccgccgc tgagcaagca gacaaccccc ttcgcctgac cagacaaatt
     3121 atcgcccgca ctaaggagct gaaatcgctg gaagaaatgg ccgctcgtgt tcccaggtat
     3181 cttcaggcaa tgctgggtta tcatagggta atgctctaca gattcgccga cgatggctct
     3241 ggtatggtaa taggggaggc caagcgatcc gacctggagt cttttctggg gcagcatttc
     3301 cccgcatctc tagtacccca gcaggcaagg cttttgtacc tcaagaatgc catccgcgtg
     3361 gtatcggact cccgcggtat tagtagcagg atcgttccag aacacgacgc ctctggagct
     3421 gcgctggatc tgtctttcgc tcacctccga tctatcagcc cttgtcattt agagtttctg
     3481 cgaaatatgg gggtcagcgc aagtatgtcc ctgagcataa taatcgatgg aacactatgg
     3541 ggcctaataa tttgccatca ttacgaacct agagcagtac caatggcaca gcgagtggcc
     3601 gccgaaatgt tcgccgattt tttgtctctt cattttactg ccgctcacca tcagcgcgga
     3661 tccgcgttta aactcgagtc tcacggcttc cctcccgagg tggaggagca ggccgccggc
     3721 accctgccca tgagctgcgc ccaggagagc ggcatggata gacaccctgc tgcttgcgcc
     3781 agcgccagga tcaacgtctt gaagttccta ttctctagaa agtataggaa cttctagtga
     3841 gcaaggtctg cagccccatc ttctccccaa gttcctcact tctcagattt tatattttct
     3901 attcctctgt gtatttatat aaaatatatt aactataacg tccccaggga ccagagcaca
     3961 gggcagtggt gctaggaggg ctcttcccag agcgaccttg tccatcctgc cacccccctc
     4021 cccccatttg cgtggaaaga aaacaaccag aagagaaaaa tcctgaactg ccaagtgcct
     4081 gcttaggagc tctgctaaga tgcccctgtt aaacttttta ggaggggtct gggtctccta
     4141 ggaccccttt tcaggcttgg gtaatagcag ccatttttag attggttctg tttatacctt
     4201 cccacccatc ttgggcaaac aaaaaaactt atattgcctg tcttccccca attctgtagg
     4261 tggaggttga tagcctagct tcctttttga gatattttca ttttgtgaaa ctctttgtaa
     4321 taaaacagct catttctata ccctcggccc tggttggttt aagttcttcc ccgtctcttg
     4381 tggggcgtca cttgcagacc tacctgcctt tgtcctccag gtaggtgcag tcctcttacc
     4441 cacaccccag aagaccactc aatgaaacat cactaaaggg acgacattta ttccttttcc
     4501 aaatgttaca gtaaatccgg gtggaagaat ggttttagca gttagaaaaa aaaaaagtac
     4561 aaatctgggg tttggccatt aaaagttatt tacaacaatg ggagggggaa aaaaaaagac
     4621 aacaagaagt tgtttcacat tacagacctc cctccctcct gaagcctaac acttgcttat
     4681 caagtcagag acatggttga tttcacaagc tggaggtttg aacttgagta agacatttat
     4741 aaaaacccag atggggcaat gtcctcccca gcccaggtgc cactaggcac agcacaagag
     4801 actaagaatt caacagggga aaggctggac actcagggtt tgggagtata agcccccact
     4861 tctggctcag ggggttgaga agcaggataa acaaaacgaa cttggcggcc gccaccgcgg
     4921 tggagctcca gcttttgttc cctttagtga gggttaattg cgcgcttggc gtaatcatgg
     4981 tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa catacgagcc
     5041 ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac attaattgcg
     5101 ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc
     5161 ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact
     5221 gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta
     5281 atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag
     5341 caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc
     5401 cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta
     5461 taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg
     5521 ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc
     5581 tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac
     5641 gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac
     5701 ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg
     5761 aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga
     5821 aggacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt
     5881 agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag
     5941 cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct
     6001 gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg
     6061 atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat
     6121 gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc
     6181 tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg
     6241 gagggcttac catctggccc cagtgctgca atgataccgc ggctgccacg ctcaccggct
     6301 ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca
     6361 actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg
     6421 ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg
     6481 tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc
     6541 cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag
     6601 ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg
     6661 ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag
     6721 tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat
     6781 agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg
     6841 atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca
     6901 gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca
     6961 aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat
     7021 tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag
     7081 aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccac
//
