LOCUS       Exported                5946 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    pCS2+ Bmi1-cherry-HA
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5946)
  AUTHORS   Pasque V, Gillich A, Garrett N, Gurdon JB
  TITLE     Histone variant macroH2A confers resistance to nuclear 
            reprogramming.
  JOURNAL   EMBO J. 2011 May 6. ():.
  PUBMED    21552206
REFERENCE   2  (bases 1 to 5946)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..5946
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     protein_bind    complement(1029..1053)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             1061..1765
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="VSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEGT
                     QTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFE
                     DGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALKG
                     EIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERAE
                     GRHSTGGMDELYK"
     primer_bind     complement(1203..1221)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1471..1490
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             1772..1852
                     /codon_start=1
                     /product="three tandem HA epitope tags"
                     /label=3xHA
                     /translation="YPYDVPDYAYPYDVPDYAYPYDVPDYA"
     promoter        complement(1861..1878)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     polyA_signal    1883..2017
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1907..1926)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1961..1980
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(2130..2150)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(2130..2148)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2169..2185)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2169..2185)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2182..2204)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2193..2209
                     /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(2217..2247)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2262..2283
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2400..2417)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2571..3159)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2651..2670)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(3330..4190)
                     /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     3953..3972
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4191..4295)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(4321..4776)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4458..4479)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4670..4689
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4850..5834
                     /label=CMV IE94 promoter
                     /note="enhancer/promoter region of simian cytomegalovirus 
                     major immediate early transcription unit IE94"
     promoter        5869..5887
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     5869..5886
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     protein_bind    5942..20
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
ORIGIN
        1 tttgtacaaa aaagcaggct ccgcggccgc ccccttcacc atgcatcgaa caaccagaat
       61 caagatcact gagctaaatc cccacttaat gtgtgtcctg tgtggagggt acttcattga
      121 tgccactacc ataatagaat gtctacattc cttctgtaaa acatgtattg tacgttactt
      181 ggagaccagc aagtattgtc ctatttgtga tgtccaggtt cacaaaacca gaccactcct
      241 gaacataagg tcagataaaa ctcttcaaga tattgtatac aaattagtcc cagggctttt
      301 caaaaatgag atgaagagaa gaagagattt ttatgcagct cacccgtcag ctgatgctgc
      361 caatggctcc aatgaagacc gaggagaagt tgcagatgag gagaagagga ttataactga
      421 tgatgagata ataagcttgt ctattgagtt ctttgatcag agcagattgg atcggaaagt
      481 aaataaagag aagcctaagg aagaggtgaa tgataaaagg tacttacgat gcccagcagc
      541 aatgactgtg atgcacttga gaaagtttct caggagtaaa atggacatac ccaatacttt
      601 ccagattgac gtcatgtatg aagaggaacc tttaaaggat tactacacgc taatggacat
      661 tgcctacatt tatacctgga gaagaaatgg cccactacct ttgaaataca gagttcggcc
      721 aacttgcaaa agaatgaaga tgagtcacca gagggatgga ctgacgaatg ctggagagct
      781 ggaaagtgac tctgggagtg acaaggccaa cagcccagcc ggaggtgttc cctccacctc
      841 ttcctgtttg cccagtccca gcactccagt gcagtctcct cacccacagt tccctcacat
      901 ttccagtacc atgaatggaa ccagcaacag ccccagtgct aaccaccaat cttcctttgc
      961 cagtagacct cgaaaatcat cactaaatgg gtcatcagca acttcatctg gtaagggtgg
     1021 gcgcgccgac ccagctttct tgtacaaagt ggttgctagc gtgagcaagg gcgaggagga
     1081 taacatggcc atcatcaagg agttcatgcg cttcaaggtg cacatggagg gctccgtgaa
     1141 cggccacgag ttcgagatcg agggcgaggg cgagggccgc ccctacgagg gcacccagac
     1201 cgccaagctg aaggtgacca agggtggccc cctgcccttc gcctgggaca tcctgtcccc
     1261 tcagttcatg tacggctcca aggcctacgt gaagcacccc gccgacatcc ccgactactt
     1321 gaagctgtcc ttccccgagg gcttcaagtg ggagcgcgtg atgaacttcg aggacggcgg
     1381 cgtggtgacc gtgacccagg actcctccct gcaggacggc gagttcatct acaaggtgaa
     1441 gctgcgcggc accaacttcc cctccgacgg ccccgtaatg cagaagaaga ccatgggctg
     1501 ggaggcctcc tccgagcgga tgtaccccga ggacggcgcc ctgaagggcg agatcaagca
     1561 gaggctgaag ctgaaggacg gcggccacta cgacgctgag gtcaagacca cctacaaggc
     1621 caagaagccc gtgcagctgc ccggcgccta caacgtcaac atcaagttgg acatcacctc
     1681 ccacaacgag gactacacca tcgtggaaca gtacgaacgc gccgagggcc gccactccac
     1741 cggcggcatg gacgagctgt acaagctcga gtacccatac gatgttccag attacgctta
     1801 tccttatgac gtacctgact atgcataccc ttatgatgta ccagactacg ctatctagaa
     1861 ctatagtgag tcgtattacg tagatccaga catgataaga tacattgatg agtttggaca
     1921 aaccacaact agaatgcagt gaaaaaaatg ctttatttgt gaaatttgtg atgctattgc
     1981 tttatttgta accattataa gctgcaataa acaagttaac aacaacaatt gcattcattt
     2041 tatgtttcag gttcaggggg aggtgtggga ggttttttaa ttcgcggccg cggcgccaat
     2101 gcattgggcc cggtacccag cttttgttcc ctttagtgag ggttaattgc gcgcttggcg
     2161 taatcatggt catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac
     2221 atacgagccg gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca
     2281 ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat
     2341 taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc
     2401 tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca
     2461 aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca
     2521 aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg
     2581 ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg
     2641 acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt
     2701 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt
     2761 tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc
     2821 tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt
     2881 gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt
     2941 agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc
     3001 tacactagaa ggacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa
     3061 agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt
     3121 tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct
     3181 acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta
     3241 tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa
     3301 agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc
     3361 tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact
     3421 acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc
     3481 tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt
     3541 ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta
     3601 agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg
     3661 tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt
     3721 acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc
     3781 agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt
     3841 actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc
     3901 tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc
     3961 gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa
     4021 ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac
     4081 tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa
     4141 aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt
     4201 tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa
     4261 tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct
     4321 aaattgtaag cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat
     4381 tttttaacca ataggccgaa atcggcaaaa tcccttataa atcaaaagaa tagaccgaga
     4441 tagggttgag tgttgttcca gtttggaaca agagtccact attaaagaac gtggactcca
     4501 acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa ccatcaccct
     4561 aatcaagttt tttggggtcg aggtgccgta aagcactaaa tcggaaccct aaagggagcc
     4621 cccgatttag agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa gggaagaaag
     4681 cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc gtaaccacca
     4741 cacccgccgc gcttaatgcg ccgctacagg gcgcgtccca ttcgccattc aggctgcgca
     4801 actgttggga agggcgatcg gtgcgggcct cttcgctatt acgccagtcg accatagcca
     4861 attcaatatg gcgtatatgg actcatgcca attcaatatg gtggatctgg acctgtgcca
     4921 attcaatatg gcgtatatgg actcgtgcca attcaatatg gtggatctgg accccagcca
     4981 attcaatatg gcggacttgg caccatgcca attcaatatg gcggacttgg cactgtgcca
     5041 actggggagg ggtctacttg gcacggtgcc aagtttgagg aggggtcttg gccctgtgcc
     5101 aagtccgcca tattgaattg gcatggtgcc aataatggcg gccatattgg ctatatgcca
     5161 ggatcaatat ataggcaata tccaatatgg ccctatgcca atatggctat tggccaggtt
     5221 caatactatg tattggccct atgccatata gtattccata tatgggtttt cctattgacg
     5281 tagatagccc ctcccaatgg gcggtcccat ataccatata tggggcttcc taataccgcc
     5341 catagccact cccccattga cgtcaatggt ctctatatat ggtctttcct attgacgtca
     5401 tatgggcggt cctattgacg tatatggcgc ctcccccatt gacgtcaatt acggtaaatg
     5461 gcccgcctgg ctcaatgccc attgacgtca ataggaccac ccaccattga cgtcaatggg
     5521 atggctcatt gcccattcat atccgttctc acgcccccta ttgacgtcaa tgacggtaaa
     5581 tggcccactt ggcagtacat caatatctat taatagtaac ttggcaagta cattactatt
     5641 ggaaggacgc cagggtacat tggcagtact cccattgacg tcaatggcgg taaatggccc
     5701 gcgatggctg ccaagtacat ccccattgac gtcaatgggg aggggcaatg acgcaaatgg
     5761 gcgttccatt gacgtaaatg ggcggtaggc gtgcctaatg ggaggtctat ataagcaatg
     5821 ctcgtttagg gaaccgccat tctgcctggg gacgtcggag caagcttgat ttaggtgaca
     5881 ctatagaata caagctactt gttctttttg caggatccca tcgattcgaa ttcaaggatc
     5941 aacaag
//
