LOCUS       Exported                5760 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  Transient expression of Pumilio1, fused to mCherry in mammalian 
            cells, under a hUBC promoter.
ACCESSION   .
VERSION     .
KEYWORDS    pUBC_hsPUM1-mCherry
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5760)
  AUTHORS   Maass PG, Barutcu AR, Shechner DM, Weiner CL, Mele M, Rinn JL
  TITLE     Spatiotemporal allele organization by allele-specific CRISPR 
            live-cell imaging (SNP-CLING).
  JOURNAL   Nat Struct Mol Biol. 2018 Jan 8. pii: 10.1038/s41594-017-0015-3. 
            doi: 10.1038/s41594-017-0015-3.
  PUBMED    29343869
REFERENCE   2  (bases 1 to 5760)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..5760
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     CDS             complement(135..995)
                     /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     758..777
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(996..1100)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     1190..1209
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     1403..1425
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     1417..1434
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     1418..1434
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2511..2530
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     regulatory      2692..2701
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             2704..2724
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2731..2757
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             3847..4557
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGREFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(3992..4010)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     4260..4279
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     4454..4477
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     primer_bind     complement(4604..4623)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4658..4677
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4734..4750)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4734..4750)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4747..4769)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4758..4774
                     /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(4782..4812)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4827..4848
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4965..4982)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5136..5724)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5216..5235)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc
       61 acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa
      121 acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta
      181 tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc
      241 ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat
      301 ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta
      361 tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt
      421 aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt
      481 ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg
      541 ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc
      601 gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc
      661 gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg
      721 cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga
      781 actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta
      841 ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct
      901 tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag
      961 ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca atattattga
     1021 agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat
     1081 aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctgacgt ctaagaaacc
     1141 attattatca tgacattaac ctataaaaat aggcgtatca cgagctatgc ggcatcagag
     1201 cagattgtac tgagagtgca ccatatgcgg tgtgaaatac cgcacagatg cgtaaggaga
     1261 aaataccgca tcaggcgcca ttcgccattc aggctgcgca actgttggga agggcgatcg
     1321 gtgcgggcct cttcgctatt acgccagctg gcgaaagggg gatgtgctgc aaggcgatta
     1381 agttgggtaa cgccagggtt ttcccagtca cgacgttgta aaacgacggc cagtgaattc
     1441 gagctcggta cacgtatgtc gaggtaggcg cgccgggttt tggcgcctcc cgcgggcgcc
     1501 cccctcctca cggcgagcgc tgccacgtca gacgaagggc gcagcgagcg tcctgatcct
     1561 tccgcccgga cgctcaggac agcggcccgc tgctcataag actcggcctt agaaccccag
     1621 tatcagcaga aggacatttt aggacgggac ttgggtgact ctagggcact ggttttcttt
     1681 ccagagagcg gaacaggcga ggaaaagtag tcccttctcg gcgattctgc ggagggatct
     1741 ccgtggggcg gtgaacgccg atgattatat aaggacgcgc cgggtgtggc acagctagtt
     1801 ccgtcgcagc cgggatttgg gtcgcggttc ttgtttgtgg atcgctgtga tcgtcacttg
     1861 gtgagtagcg ggctgctggg ctggccgggg ctttcgtggc cgccgggccg ctcggtggga
     1921 cggaagcgtg tggagagacc gccaagggct gtagtctggg tccgcgagca aggttgccct
     1981 gaactggggg ttggggggag cgcagcaaaa tggcggctgt tcccgagtct tgaatggaag
     2041 acgcttgtga ggcgggctgt gaggtcgttg aaacaaggtg gggggcatgg tgggcggcaa
     2101 gaacccaagg tcttgaggcc ttcgctaatg cgggaaagct cttattcggg tgagatgggc
     2161 tggggcacca tctggggacc ctgacgtgaa gtttgtcact gactggagaa ctcggtttgt
     2221 cgtctgttgc gggggcggca gttatggcgg tgccgttggg cagtgcaccc gtacctttgg
     2281 gagcgcgcgc cctcgtcgtg tcgtgacgtc acccgttctg ttggcttata atgcagggtg
     2341 gggccacctg ccggtaggtg tgcggtaggc ttttctccgt cgcaggacgc agggttcggg
     2401 cctagggtag gctctcctga atcgacaggc gccggacctc tggtgagggg agggataagt
     2461 gaggcgtcag tttctttggt cggttttatg tacctatctt cttaagtagc tgaagctccg
     2521 gttttgaact atgcgctcgg ggttggcgag tgtgttttgt gaagtttttt aggcaccttt
     2581 tgaaatgtaa tcatttgggt caatatgtaa ttttcagtgt tagactagta aattgtccgc
     2641 taaattctgg ccgtttttgg cttttttgtt agacctggag gtaccgccac cgccaccatg
     2701 gctccaaaga agaagcgtaa ggtaggcagc tacccatacg atgttccaga ttacgctggc
     2761 agcggtcggt cccggctctt ggaggacttt cgaaacaatc gatatcctaa cttgcagctt
     2821 cgcgagatcg ctggtcatat tatggaattc tctcaggatc aacacgggtc tagattcatt
     2881 cagctcaagc tggagagagc cacacctgcc gaacggcagc tcgtgtttaa tgagatcctc
     2941 caggctgcct atcaactgat ggtggacgtg ttcggcaatt acgtcatcca gaagttcttt
     3001 gaatttggta gtctcgagca gaagctggcc cttgcagaga gaatccgcgg ccatgtactg
     3061 tcccttgccc tgcagatgta cgggtgccgc gtaatccaga aggcactgga atttataccg
     3121 tccgaccagc agaatgagat ggtgcgggaa ctcgatggtc atgtgttgaa atgcgtaaaa
     3181 gatcagaacg gtaaccacgt ggttcagaaa tgtatcgagt gtgtccagcc acagagcttg
     3241 caattcatca tagatgcctt taagggccag gttttcgctc tgagcacgca cccctacgga
     3301 tgccgggtca ttcagcggat tcttgaacat tgtctgccag accagactct gcctatcctg
     3361 gaggagctcc accaacacac cgaacagttg gtccaggatc aatatggtaa ttacgttatt
     3421 cagcatgtgc tggaacatgg gcggcctgag gacaagtcca aaattgtagc tgagattcga
     3481 ggcaatgtgc tcgttctttc ccagcacaag ttcgcttcta acgtcgtgga gaaatgtgtg
     3541 actcacgcca gccggactga gcgcgcggtg ctcattgatg aagtgtgtac catgaacgat
     3601 gggccacata gcgcacttta cacaatgatg aaagaccagt acgcaaacta cgtggttcag
     3661 aagatgatag acgtcgctga accggggcag cgcaaaattg tcatgcacaa gatccgccca
     3721 cacatagcta ctcttcggaa atacacctat ggcaagcaca tcctcgcaaa gctggagaag
     3781 tattacatga aaaacggggt ggatctggga ggatccaagg gctcgggctc gacctcgggc
     3841 tccggaatgg tgagcaaggg cgaggaggac aacatggcca tcatcaagga gttcatgcgc
     3901 ttcaaggtgc acatggaggg ctccgtgaac ggccgcgagt tcgagatcga gggcgagggc
     3961 gagggccgcc cctacgaggg cacccagacc gccaagctga aggtgaccaa gggcggcccc
     4021 ctgcccttcg cctgggacat cctgtcccct cagttcatgt acggctccaa ggcctacgtg
     4081 aagcaccccg ccgacatccc cgactacttg aagctgtcct tccccgaggg cttcaagtgg
     4141 gagcgcgtga tgaacttcga ggacggcggc gtggtgaccg tgacccagga ctcctccctg
     4201 caggacggcg agttcatcta caaggtgaag ctgcgcggca ccaacttccc ctccgacggc
     4261 cccgtaatgc agaagaagac catgggctgg gaggcctcct ccgagcggat gtaccccgag
     4321 gacggcgccc tgaagggcga gatcaagcag aggctgaagc tgaaggacgg cggccactac
     4381 gacgccgagg tcaagaccac ctacaaggcc aagaagcccg tgcagctgcc cggcgcctac
     4441 aacgtcaaca tcaagctgga catcacctcc cacaacgagg actacaccat cgtggaacag
     4501 tacgagcgcg ccgagggccg ccactccacc ggcggcatgg acgagctgta caagtaatga
     4561 taagtcgact tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat
     4621 ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat
     4681 gtatcttatt cgactgttta aacctgcagg catgcaagct tggcgtaatc atggtcatag
     4741 ctgtttcctg tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagc
     4801 ataaagtgta aagcctgggg tgcctaatga gtgagctaac tcacattaat tgcgttgcgc
     4861 tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa
     4921 cgcgcgggga gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg
     4981 ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg
     5041 ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag
     5101 gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac
     5161 gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga
     5221 taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt
     5281 accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc
     5341 tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc
     5401 cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta
     5461 agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat
     5521 gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca
     5581 gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct
     5641 tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt
     5701 acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct
//
