LOCUS       sequence-200613-        8100 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Donor for integration at the human ROSA26 locus.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8100)
  AUTHORS   Okumura M, Natsume T, Kanemaki MT, Kiyomitsu T
  TITLE     Dynein-Dynactin-NuMA clusters generate cortical spindle-pulling 
            forces as a multi-arm ensemble.
  JOURNAL   Elife. 2018 May 31;7. pii: 36559. doi: 10.7554/eLife.36559.
  PUBMED    29848445
REFERENCE   2  (bases 1 to 8100)
  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: "Elife."; 
            date: "2018-05-31"; volume: "7. pii: 36559. doi"; pages: " 
            10.7554/eLife.36559"
COMMENT     Sequence Label: pMK247 (hROSA26 CMV-MCS-Hygro)
FEATURES             Location/Qualifiers
     source          1..8100
                     /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"
     promoter        626..644
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     enhancer        1600..1979
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1980..2183
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2134..2154
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     promoter        2228..2246
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     intron          2339..2568
                     /label=chimeric intron
                     /note="chimera between introns from adenovirus and 
                     immunoglobulin heavy chain genes"
     primer_bind     2551..2570
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     misc_feature    join(2639..3213,3214..3216,3217..3225)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(2806..2823)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     CDS             3252..4271
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGYV
                     LRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPET
                     ELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQTV
                     MDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFGD
                     SQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGNF
                     DDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE"
     CDS             3252..4271
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGYV
                     LRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPET
                     ELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQTV
                     MDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFGD
                     SQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGNF
                     DDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE"
     polyA_signal    4585..4808
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(5912..5932)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(5912..5930)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(5951..5967)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5951..5967)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5964..5986)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5975..5991
                     /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(5999..6005,6006..6023,6024..6029))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6044..6065
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6182..6199)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6353..6941)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6433..6452)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7112..7903,7904..7972))
                     /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     7735..7754
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7973..8077)
                     /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 tggagctcgg
      661 aactctggtg ggaggggagg tgcggtacac tggggggatg ggtggctagg ggggccgtct
      721 ggtggcttgc gggggttgcc tttcccgtgg gaagtcggga acataatgtt tgttacgttg
      781 ggagggaaag gggtggctgg atgcaggcgg gagggaggcc cgccctgcgg caaccggagg
      841 gggagggaga agggagcgga aaatgctcga aaccggacgg agccattgct ctcgcagagg
      901 gaggagcgct tccggctagc ctcttgtcgc cgattggccg tttctcctcc cgccgtgtgt
      961 gaaaacacaa atggcgtatt ctggttggag taaagctcct gtcagttacg ccgtcgggag
     1021 tacgcagccg cttagcgact ctcgcgttgc cccctgggtg gggcgggtag gtaggtgggg
     1081 tgtagagatg ctgggtgtgc gggcgcggcc ggcctcctgc ggcgggaggg gagggtcagt
     1141 gaaatcggct ctggcgcggg cgtcctccca ccctcccctt ccttcggggg agtcggttta
     1201 cccgccgcct gcttgtcttc gacacctgat tggctgtcga agctgtggga ccgggccctt
     1261 gctactggct cgagtctcac atgagcgaaa ccactgcgcg gggcgcgggg gtggcgggga
     1321 ggcgggcgtt ggtacggtcc tccccgaggc cgagcgccgc agtgtctggc cccgcgcccc
     1381 tgcgcaacgt ggcaggaagc gcgcgctgga ggcgggggcg ggctgccggc cgagacttct
     1441 ggatggcggc ggccgcggct ccgccccggg ttcccaccgc ctgaagggcg agacaagccc
     1501 gacctgctac aggcactcgt gggggtgggg gaggagcggg ggtcggtccg gctggtttgt
     1561 gggtgggagg cgcttgttct ccaaaaaccg gcgcgcgttg acattgatta ttgactagtt
     1621 attaatagta atcaattacg gggtcattag ttcatagccc atatatggag ttccgcgtta
     1681 cataacttac ggtaaatggc ccgcctggct gaccgcccaa cgacccccgc ccattgacgt
     1741 caataatgac gtatgttccc atagtaacgc caatagggac tttccattga cgtcaatggg
     1801 tggactattt acggtaaact gcccacttgg cagtacatca agtgtatcat atgccaagta
     1861 cgccccctat tgacgtcaat gacggtaaat ggcccgcctg gcattatgcc cagtacatga
     1921 ccttatggga ctttcctact tggcagtaca tctacgtatt agtcatcgct attaccatgg
     1981 tgatgcggtt ttggcagtac atcaatgggc gtggatagcg gtttgactca cggggatttc
     2041 caagtctcca ccccattgac gtcaatggga gtttgttttg gcaccaaaat caacgggact
     2101 ttccaaaatg tcgtaacaac tccgccccat tgacgcaaat gggcggtagg cgtgtacggt
     2161 gggaggtcta tataagcaga gctctctggc taactagaga acccactgct tactggctta
     2221 tcgaaattaa tacgactcac tatagggaga cccaagcttg gtaccgagct cggatccact
     2281 agtaacggcc gccagtgtgc tggaattaat tcgctgtctg cgagggccag ctgttggggt
     2341 gagtactccc tctcaaaagc gggcatgact tctgcgctaa gattgtcagt ttccaaaaac
     2401 gaggaggatt tgatattcac ctggcccgcg gtgatgcctt tgagggtggc cgcgtccatc
     2461 tggtcagaaa agacaatctt tttgttgtca agcttgaggt gtggcaggct tgagatctgg
     2521 ccatacactt gagtgacaat gacatccact ttgcctttct ctccacaggt gtccactccc
     2581 aggtccaact gcaggtcgat cgagcatgca tctagggcgg ccgcactaga ggaattcgcc
     2641 cctctccctc ccccccccct aacgttactg gccgaagccg cttggaataa ggccggtgtg
     2701 tgtttgtcta tatgtgattt tccaccatat tgccgtcttt tggcaatgtg agggcccgga
     2761 aacctggccc tgtcttcttg acgagcattc ctaggggtct ttcccctctc gccaaaggaa
     2821 tgcaaggtct gttgaatgtc gtgaaggaag cagttcctct ggaagcttct tgaagacaaa
     2881 caacgtctgt agcgaccctt tgcaggcagc ggaacccccc acctggcgac aggtgcctct
     2941 gcggccaaaa gccacgtgta taagatacac ctgcaaaggc ggcacaaccc cagtgccacg
     3001 ttgtgagttg gatagttgtg gaaagagtca aatggctctc ctcaagcgta gtcaacaagg
     3061 ggctgaagga tgcccagaag gtaccccatt gtatgggaat ctgatctggg gcctcggtgc
     3121 acatgcttta catgtgttta gtcgaggtta aaaaagctct aggccccccg aaccacgggg
     3181 acgtggtttt cctttgaaaa acacgatgat aagcttgcca caaccccgta ccaaagatgg
     3241 atagatccgg aaagcctgaa ctcaccgcga cgtctgtcga gaagtttctg atcgaaaagt
     3301 tcgacagcgt ctccgacctg atgcagctct cggagggcga agaatctcgt gctttcagct
     3361 tcgatgtagg agggcgtgga tatgtcctgc gggtaaatag ctgcgccgat ggtttctaca
     3421 aagatcgtta tgtttatcgg cactttgcat cggccgcgct cccgattccg gaagtgcttg
     3481 acattgggga attcagcgag agcctgacct attgcatctc ccgccgtgca cagggtgtca
     3541 cgttgcaaga cctgcctgaa accgaactgc ccgctgttct gcagccggtc gcggaggcca
     3601 tggatgcgat cgctgcggcc gatcttagcc agacgagcgg gttcggccca ttcggaccgc
     3661 aaggaatcgg tcaatacact acatggcgtg atttcatatg cgcgattgct gatccccatg
     3721 tgtatcactg gcaaactgtg atggacgaca ccgtcagtgc gtccgtcgcg caggctctcg
     3781 atgagctgat gctttgggcc gaggactgcc ccgaagtccg gcacctcgtg cacgcggatt
     3841 tcggctccaa caatgtcctg acggacaatg gccgcataac agcggtcatt gactggagcg
     3901 aggcgatgtt cggggattcc caatacgagg tcgccaacat cttcttctgg aggccgtggt
     3961 tggcttgtat ggagcagcag acgcgctact tcgagcggag gcatccggag cttgcaggat
     4021 cgccgcggct ccgggcgtat atgctccgca ttggtcttga ccaactctat cagagcttgg
     4081 ttgacggcaa tttcgatgat gcagcttggg cgcagggtcg atgcgacgca atcgtccgat
     4141 ccggagccgg gactgtcggg cgtacacaaa tcgcccgcag aagcgcggcc gtctggaccg
     4201 atggctgtgt agaagtactc gccgatagtg gaaaccgacg ccccagcact cgtccgaggg
     4261 caaaggaata gagtagatgc cgaccgaaca agagctgatt tcgagaacgc ctcagccagc
     4321 aactcgcgcg agcctagcaa ggcaaatgcg agagaacggc cttacgcttg gtggcacagt
     4381 tctcgtccac agttcgctaa gctcgctcgg ctgggtcgcg ggagggccgg tcgcagtgat
     4441 tcaggccctt ctggattgtg ttggtcccca gggcacgatt gtcatgccca cgcactcggg
     4501 tgatctgact gatcccgcag attggagatc gccgcccgtg cctgccgatt gggtgcagat
     4561 ctagagctcg ctgatcagcc tcgactgtgc ctctagttgc cagccatctg ttgtttgccc
     4621 ctcccccgtg ccttccttga ccctggaagg tgccactccc actgtccttt cctaataaaa
     4681 tgaggaaatt gcatcgcatt gtctgagtag gtgtcattct attctggggg gtggggtggg
     4741 gcaggacagc aagggggagg attgggaaga caatagcagg catgctgggg atgcggtggg
     4801 ctctatggct tctgaggcgg aaagaaccag ctggggctcg atgagggagc tgcggtggag
     4861 gaggtggaga gaaggccgca cccttctggg cagggggagg ggagtgccgc aataccttta
     4921 tgggagttct ctgctgcctc ccgtcttgta aggaccgccc tgggcctgga agaagccctc
     4981 cctcctttcc tcctcgcgtg atctcgtcat cgcctccatg tcgagtcgct tctcgattat
     5041 gggcgggatt cttttgccta ggcttaaggg gctaacttgg tccctgggcg ttgccctgca
     5101 ggggagtgag cagctgtaag atttgagggg cgactccgat tagtttatct tcccacggac
     5161 tagagttggt gtcgaggtta ttgtaataag ggtggggtag ggaaatggag cttagtcatt
     5221 cacctggggc tgattttatg caacgagact gcggattatc actacttatc atttttggag
     5281 catttttcta gagacagaca taaagcatga tcacctgagt tttataccat ttgagaccct
     5341 tgctgcacca ccaaagtgta gcatcaggtt aaatcttaat agaaaaattt tagcttttgc
     5401 ttgagaaacc agtgcttccc tccctcaccc tctctcccca ggctctctac ccctttgcat
     5461 ccctaccagg catcttagca actctcactc atacttgatc ccattttcca tttgttgtac
     5521 ttgctcctct agtattcaga catagcacta gctttctccc tctcttgatc ttgggtagcc
     5581 tggtgtctcg cgaaaccaga cagattggtt ccaccacaaa ttaaggcttg agctggggct
     5641 tgactcttac ccagcagtgc ttttattcct ccctagttca cgttcttaaa tgtttatctt
     5701 gattttcatt ttatcctttt tccttagctg ggattctgtc cctgaccgtc ttcacagtcc
     5761 aggtgatctt gactactgct ttacagagaa ttggatctga ggttaggcaa catctccctt
     5821 tttcttcctc taaatacctc tcatttctgt tcttaccagt tagtaactga tctcagatgc
     5881 ctgtgtgata gctggtaccc agcttttgtt ccctttagtg agggttaatt gcgcgcttgg
     5941 cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta tccgctcaca attccacaca
     6001 acatacgagc cggaagcata aagtgtaaag cctggggtgc ctaatgagtg agctaactca
     6061 cattaattgc gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc
     6121 attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt
     6181 cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact
     6241 caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag
     6301 caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata
     6361 ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc
     6421 cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg
     6481 ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc
     6541 tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg
     6601 gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc
     6661 ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga
     6721 ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg
     6781 gctacactag aaggacagta tttggtatct gcgctctgct gaagccagtt accttcggaa
     6841 aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg
     6901 tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt
     6961 ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat
     7021 tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct
     7081 aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta
     7141 tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa
     7201 ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac
     7261 gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa
     7321 gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag
     7381 taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg
     7441 tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag
     7501 ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg
     7561 tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc
     7621 ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat
     7681 tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata
     7741 ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa
     7801 aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca
     7861 actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc
     7921 aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc
     7981 tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg
     8041 aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac
//
