LOCUS       sequence-238910-        7092 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Targeting vector for mouse Birc6 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 7092)
  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 7092)
  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-Birc6-tdiRFP670-PEST
FEATURES             Location/Qualifiers
     source          1..7092
                     /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             1636..1689
                     /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(1690..1723)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             1732..1752
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             3715..3834
                     /codon_start=1
                     /product="PEST degradation sequence from mouse ornithine 
                     decarboxylase"
                     /label=hPEST
                     /note="human codon-optimized"
                     /translation="SHGFPPEVEEQAAGTLPMSCAQESGMDRHPAACASARINV"
     protein_bind    3837..3870
                     /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(4905..4925)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(4905..4923)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(4944..4960)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4944..4960)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4957..4979)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4968..4984
                     /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(4992..4998,4999..5016,5017..5022))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5037..5058
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5175..5192)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5346..5934)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(join(6105..6896,6897..6965))
                     /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     6728..6747
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6966..7056)
                     /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 gaagggcggt cggtgcgggc ctcttcgcta ttacgccagc tggcgaaagg
      541 gggatgtgct gcaaggcgat taagttgggt aacgccaggg ttttcccagt cacgacgttg
      601 taaaacgacg gccagtgagc gcgcgtaata cgactcacta tagggcgaat tgggtacccc
      661 ttgctcgtgc cagagaagtg ctgcactatt caatcaacac ccaatctggg tatattttag
      721 tttgttcagt tgtccttttc agagttggca tctgtgacag cacaaggaat tactgtcttt
      781 aatttaaaag aatatgtata tcaaactatt gatgtttgta agtcattgtt gatgctgatc
      841 ctgtggaact gtactgtgtt cttgaggagg tggagtttag tcttttgtag ctggatagaa
      901 tgtaactctg agcttttctt ttcttttttt tttttctttt ttcttttttt tggttttttg
      961 agacagggtt tctctgtgta gccctggctg tcctggaact cactttgtag accaggctgg
     1021 ccacgaactc agaaatctgc ctgcctctgc ctcccaagtg ctgggattaa aggcgccacc
     1081 acacccagct tactctgagc ttttcaacat ctggaattaa tttctatgtt taggaatttt
     1141 gcctgcatgt atatgcatat gggggtggtg tgagccctgg aacttgagtt acagacaggt
     1201 gtgaattacc atgtgggtgc tgggaattga accagggtcc tcatgaaaaa tagctagtgc
     1261 tcttaacctc tgaaccatct ctctagcccc tagtttgggg atccttgaga gttttgttta
     1321 ctgtgtttta aatggctgcc ataagaagac agtaaagatt tgtactcgat actgaagtat
     1381 gttaaaacca agttgtctca gtgataggca agaatcacac tgttcacact tctgatatta
     1441 gcgattttct ctacctagcg ccacactgct cagctccgag aagagctact gaaacttccc
     1501 tgccctgaag gcttagatcc ggacattgag gatgcctccc cggtgtgcag ggccacagca
     1561 ggtgctgaag acactctaac ccacgaccat gtcaatccca gcagcagcaa agacctcccc
     1621 agtgacttcc aattagaggg aagaggtagc ctgctaactt gcggggacgt agaggaaaat
     1681 cctggtccca taacttcgta tagcatacat tatacgaagt tatcaatggg cccaaaaaag
     1741 aaaagaaaag ttgaattcat ggccagaaaa gtggacctga ccagttgtga ccgagagccg
     1801 atacacattc ctggctctat ccagccatgt gggtgtttgc tggcctgcga tgcccaggca
     1861 gtcaggatta ctcgaattac cgaaaatgcg ggtgccttct ttggccgtga aacgccccgt
     1921 gtcggcgaac tcttagccga ttatttcgga gaaaccgagg ctcacgcact taggaacgca
     1981 ctggcgcaat caagcgatcc caaacgccct gctctcatct ttgggtggag agatggactg
     2041 acaggccgta ctttcgacat aagccttcac cggcatgacg gaaccagtat aattgagttt
     2101 gaaccggccg ccgctgagca agcagacaac ccccttcgcc tgaccagaca aattatcgcc
     2161 cgcactaagg agctgaaatc gctggaagaa atggccgctc gtgttcccag gtatcttcag
     2221 gcaatgctgg gttatcatag ggtaatgctc tacagattcg ccgacgatgg ctctggtatg
     2281 gtaatagggg aggccaagcg atccgacctg gagtcttttc tggggcagca tttccccgca
     2341 tctctagtac cccagcaggc aaggcttttg tacctcaaga atgccatccg cgtggtatcg
     2401 gactcccgcg gtattagtag caggatcgtt ccagaacacg acgcctctgg agctgcgctg
     2461 gatctgtctt tcgctcacct ccgatctatc agcccttgtc atttagagtt tctgcgaaat
     2521 atgggggtca gcgcaagtat gtccctgagc ataataatcg atggaacact atggggccta
     2581 ataatttgcc atcattacga acctagagca gtaccaatgg cacagcgagt ggccgccgaa
     2641 atgttcgccg attttttgtc tcttcatttt actgccgctc accatcagcg cggccacggg
     2701 actggctcga caggttctgg cagtagtggg actgcttcat cagaaaataa taacatggcc
     2761 atggccagaa aagtggacct gaccagttgt gaccgagagc cgatacacat tcctggctct
     2821 atccagccat gtgggtgttt gctggcctgc gatgcccagg cagtcaggat tactcgaatt
     2881 accgaaaatg cgggtgcctt ctttggccgt gaaacgcccc gtgtcggcga actcttagcc
     2941 gattatttcg gagaaaccga ggctcacgca cttaggaacg cactggcgca atcaagcgat
     3001 cccaaacgcc ctgctctcat ctttgggtgg agagatggac tgacaggccg tactttcgac
     3061 ataagccttc accggcatga cggaaccagt ataattgagt ttgaaccggc cgccgctgag
     3121 caagcagaca acccccttcg cctgaccaga caaattatcg cccgcactaa ggagctgaaa
     3181 tcgctggaag aaatggccgc tcgtgttccc aggtatcttc aggcaatgct gggttatcat
     3241 agggtaatgc tctacagatt cgccgacgat ggctctggta tggtaatagg ggaggccaag
     3301 cgatccgacc tggagtcttt tctggggcag catttccccg catctctagt accccagcag
     3361 gcaaggcttt tgtacctcaa gaatgccatc cgcgtggtat cggactcccg cggtattagt
     3421 agcaggatcg ttccagaaca cgacgcctct ggagctgcgc tggatctgtc tttcgctcac
     3481 ctccgatcta tcagcccttg tcatttagag tttctgcgaa atatgggggt cagcgcaagt
     3541 atgtccctga gcataataat cgatggaaca ctatggggcc taataatttg ccatcattac
     3601 gaacctagag cagtaccaat ggcacagcga gtggccgccg aaatgttcgc cgattttttg
     3661 tctcttcatt ttactgccgc tcaccatcag cgcggatccg cgtttaaact cgagtctcac
     3721 ggcttccctc ccgaggtgga ggagcaggcc gccggcaccc tgcccatgag ctgcgcccag
     3781 gagagcggca tggatagaca ccctgctgct tgcgccagcg ccaggatcaa cgtcttgaag
     3841 ttcctattct ctagaaagta taggaacttc tagtgagctg cattgatgtg gatttcgtag
     3901 acacaaaggc ttcgaagcac aagccaaata tgtcaatatt tgtatgtaag aaactaatta
     3961 tgtaataggt aatgaaactg aaactatact atgcccttaa ggagatccag tttaattcaa
     4021 ggtgatcttt gatttacctg tacaagagtg taaacttttt tgtgctttta tttttcaatt
     4081 gtgagaacca ctgattggta tgttcaacaa atttgtgtat acaaaaaaat ggataaatca
     4141 ctgatctata agggaaacta ccttaggaaa gaatggttac tgaatgttta tctttttttt
     4201 ttttttttac tgtagagtga gaggttgtta acaaagaata tatattggtc attcttacaa
     4261 ctactattta aagtcagcaa cttttcactg aatttgatag attttgtttg gccatatctt
     4321 catgctcata tttgatttct gaagacttcc tacatacact ttaataaaag ttaaacagac
     4381 atagcccctc ttttttgatt tactggtaca tttttaagta ataaatctgc cataaaatgc
     4441 actatagctg gagacttgca cttgtatgga tgaatttatt acattcaaca tatttaattt
     4501 tatgccttct aagatgcaga aaaaaataaa tgaacatgat tttattctat gccaaacatt
     4561 tgggcctctg aatgtatcca ttatttgaat ttactatatg aaaaggtatg gtcaattgga
     4621 aaagtcgctt taaactgttt tcctcaaggg ctcctcttat tcctggactg tgtgggtttg
     4681 ttcctgaagt cacatgtatg tatcaacatg gaggctctgt agaggagaga ggctgggcct
     4741 ctctggtgct gttacagtac gttctgtacc tgccatacag gctcgttttc atgcaaattc
     4801 ttcctagagc caataaataa aaatgtaggt aaattagtat gtcttgtttc taaaatagtt
     4861 gccatggtgt gcaatggtat ttctacgagc tccagctttt gttcccttta gtgagggtta
     4921 attgcgcgct tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc
     4981 acaattccac acaacatacg agccggaagc ataaagtgta aagcctgggg tgcctaatga
     5041 gtgagctaac tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg
     5101 tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg
     5161 cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg
     5221 gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga
     5281 aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg
     5341 gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag
     5401 aggtggcgaa acccgacagg actataaaga taccaggcgt ttcctcctgg aagctccctc
     5461 gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg
     5521 ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt
     5581 cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc
     5641 ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc
     5701 actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg
     5761 tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca
     5821 gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc
     5881 ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat
     5941 cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt
     6001 ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt
     6061 tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc
     6121 agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc
     6181 gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata
     6241 ccgcggctgc cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg
     6301 gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc
     6361 cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct
     6421 acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa
     6481 cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt
     6541 cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca
     6601 ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac
     6661 tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca
     6721 atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt
     6781 tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc
     6841 actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca
     6901 aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata
     6961 ctcatactct tccttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg
     7021 gatacatatt tgaatgtatt tagaaaaata aacaaatagg gattccgcgc acatttcccc
     7081 gaaaagtgcc ac
//
