LOCUS       Exported                8879 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  Expresses sgRNA for telomere repeats with fluorescent indicator BFP 
            and puromycin selection marker.
ACCESSION   .
VERSION     .
KEYWORDS    pLV-sgTel
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8879)
  AUTHORS   Shin Y, Chang YC, Lee DSW, Berry J, Sanders DW, Ronceray P, Wingreen
            NS, Haataja M, Brangwynne CP
  TITLE     Liquid Nuclear Condensates Mechanically Sense and Restructure the 
            Genome.
  JOURNAL   Cell. 2018 Nov 29;175(6):1481-1491.e13. doi: 
            10.1016/j.cell.2018.10.057.
  PUBMED    30500535
REFERENCE   2  (bases 1 to 8879)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..8879
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        382..580
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             598..778
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    825..950
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1447..1680
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1865..1909
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
     misc_feature    2207..2324
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     protein_bind    2577..2610
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     primer_bind     2614..2634
                     /label=mU6-F
                     /note="Mouse U6 promoter, forward primer"
     promoter        2881..3092
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     intron          3093..4031
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     3988..4008
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     CDS             4052..4648
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(4052..4071)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4548..4568
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     CDS             4658..4711
                     /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"
     CDS             4724..5422
                     /codon_start=1
                     /product="monomeric blue fluorescent protein"
                     /label=TagBFP
                     /note="mammalian codon-optimized"
                     /translation="SELIKENMHMKLYMEGTVDNHHFKCTSEGEGKPYEGTQTMRIKVV
                     EGGPLPFAFDILATSFLYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLTAT
                     QDTSLQDGCLIYNVKIRGVNFTSNGPVMQKKTLGWEAFTETLYPADGGLEGRNDMALKL
                     VGGSHLIANIKTTYRSKKPAKNLKMPGVYYVDYRLERIKEANNETYVEQHEVAVARYCD
                     LPSKLGHKLN"
     protein_bind    5442..5475
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     misc_feature    5531..6119
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(5584..5604)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(6002..6013)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(6122..6138)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6123..6139)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             6648..6828
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     rep_origin      complement(6890..7478)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6970..6989)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(7649..8509)
                     /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     8272..8291
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8510..8614)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(8689..8708)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag cgcgttttgc ctgtactggg
      601 tctctctggt tagaccagat ctgagcctgg gagctctctg gctaactagg gaacccactg
      661 cttaagcctc aataaagctt gccttgagtg cttcaagtag tgtgtgcccg tctgttgtgt
      721 gactctggta actagagatc cctcagaccc ttttagtcag tgtggaaaat ctctagcagt
      781 ggcgcccgaa cagggacttg aaagcgaaag ggaaaccaga ggagctctct cgacgcagga
      841 ctcggcttgc tgaagcgcgc acggcaagag gcgaggggcg gcgactggtg agtacgccaa
      901 aaattttgac tagcggaggc tagaaggaga gagatgggtg cgagagcgtc agtattaagc
      961 gggggagaat tagatcgcga tgggaaaaaa ttcggttaag gccaggggga aagaaaaaat
     1021 ataaattaaa acatatagta tgggcaagca gggagctaga acgattcgca gttaatcctg
     1081 gcctgttaga aacatcagaa ggctgtagac aaatactggg acagctacaa ccatcccttc
     1141 agacaggatc agaagaactt agatcattat ataatacagt agcaaccctc tattgtgtgc
     1201 atcaaaggat agagataaaa gacaccaagg aagctttaga caagatagag gaagagcaaa
     1261 acaaaagtaa gaccaccgca cagcaagcgg ccggccgctg atcttcagac ctggaggagg
     1321 agatatgagg gacaattgga gaagtgaatt atataaatat aaagtagtaa aaattgaacc
     1381 attaggagta gcacccacca aggcaaagag aagagtggtg cagagagaaa aaagagcagt
     1441 gggaatagga gctttgttcc ttgggttctt gggagcagca ggaagcacta tgggcgcagc
     1501 gtcaatgacg ctgacggtac aggccagaca attattgtct ggtatagtgc agcagcagaa
     1561 caatttgctg agggctattg aggcgcaaca gcatctgttg caactcacag tctggggcat
     1621 caagcagctc caggcaagaa tcctggctgt ggaaagatac ctaaaggatc aacagctcct
     1681 ggggatttgg ggttgctctg gaaaactcat ttgcaccact gctgtgcctt ggaatgctag
     1741 ttggagtaat aaatctctgg aacagatttg gaatcacacg acctggatgg agtgggacag
     1801 agaaattaac aattacacaa gcttaataca ctccttaatt gaagaatcgc aaaaccagca
     1861 agaaaagaat gaacaagaat tattggaatt agataaatgg gcaagtttgt ggaattggtt
     1921 taacataaca aattggctgt ggtatataaa attattcata atgatagtag gaggcttggt
     1981 aggtttaaga atagtttttg ctgtactttc tatagtgaat agagttaggc agggatattc
     2041 accattatcg tttcagaccc acctcccaac cccgagggga cccgacaggc ccgaaggaat
     2101 agaagaagaa ggtggagaga gagacagaga cagatccatt cgattagtga acggatcggc
     2161 actgcgtgcg ccaattctgc agacaaatgg cagtattcat ccacaatttt aaaagaaaag
     2221 gggggattgg ggggtacagt gcaggggaaa gaatagtaga cataatagca acagacatac
     2281 aaactaaaga attacaaaaa caaattacaa aaattcaaaa ttttcgggtt tattacaggg
     2341 acagcagaga tccagtttgg ttagtaccgg gcccgctcta gagatccgac gcgccatctc
     2401 taggcccgcg ccggccccct cgcacggact tgtgggagaa gctcggctac tcccctgccc
     2461 cggttaattt gcatataata tttcctagta actatagagg cttaatgtgc gataaaagac
     2521 agataatctg ttctttttaa tactagctac attttacatg ataggcttgg atttctataa
     2581 cttcgtatag catacattat acgaagttat aaacagcaca aaaggaaact caccctaact
     2641 gtaaagtaat tgtgtgtttt gagactataa gtatcccttg gagaaccacc ttgttggtta
     2701 gggttagggt tagggttagt ttaagagcta tgctggaaac agcatagcaa gtttaaataa
     2761 ggctagtccg ttatcaactt gaaaaagtgg caccgagtcg gtgctttttt tctcgagtac
     2821 taggatccat taggcggccg cgtggataac cgtattaccg ccatgcatgt gcccgtcagt
     2881 gggcagagcg cacatcgccc acagtccccg agaagttggg gggaggggtc ggcaattgaa
     2941 ccggtgccta gagaaggtgg cgcggggtaa actgggaaag tgatgtcgtg tactggctcc
     3001 gcctttttcc cgagggtggg ggagaaccgt atataagtgc agtagtcgcc gtgaacgttc
     3061 tttttcgcaa cgggtttgcc gccagaacac aggtaagtgc cgtgtgtggt tcccgcgggc
     3121 ctggcctctt tacgggttat ggcccttgcg tgccttgaat tacttccacc tggctgcagt
     3181 acgtgattct tgatcccgag cttcgggttg gaagtgggtg ggagagttcg aggccttgcg
     3241 cttaaggagc cccttcgcct cgtgcttgag ttgaggcctg gcctgggcgc tggggccgcc
     3301 gcgtgcgaat ctggtggcac cttcgcgcct gtctcgctgc tttcgataag tctctagcca
     3361 tttaaaattt ttgatgacct gctgcgacgc tttttttctg gcaagatagt cttgtaaatg
     3421 cgggccaaga tctgcacact ggtatttcgg tttttggggc cgcgggcggc gacggggccc
     3481 gtgcgtccca gcgcacatgt tcggcgaggc ggggcctgcg agcgcggcca ccgagaatcg
     3541 gacgggggta gtctcaagct ggccggcctg ctctggtgcc tggcctcgcg ccgccgtgta
     3601 tcgccccgcc ctgggcggca aggctggccc ggtcggcacc agttgcgtga gcggaaagat
     3661 ggccgcttcc cggccctgct gcagggagct caaaatggag gacgcggcgc tcgggagagc
     3721 gggcgggtga gtcacccaca caaaggaaaa gggcctttcc gtcctcagcc gtcgcttcat
     3781 gtgactccac ggagtaccgg gcgccgtcca ggcacctcga ttagttctcg cgcttttgga
     3841 gtacgtcgtc tttaggttgg ggggaggggt tttatgcgat ggagtttccc cacactgagt
     3901 gggtggagac tgaagttagg ccagcttggc acttgatgta attctccttg gaatttgccc
     3961 tttttgagtt tggatcttgg ttcattctca agcctcagac agtggttcaa agtttttttc
     4021 ttccatttca ggtgtcgtga gctagcccac catgaccgag tacaagccca cggtgcgcct
     4081 cgccacccgc gacgacgtcc cccgggccgt acgcaccctc gccgccgcgt tcgccgacta
     4141 ccccgccacg cgccacaccg tcgacccgga ccgccacatc gagcgggtca ccgagctgca
     4201 agaactcttc ctcacgcgcg tcgggctcga catcggcaag gtgtgggtcg cggacgacgg
     4261 cgccgcggtg gcggtctgga ccacgccgga gagcgtcgaa gcgggggcgg tgttcgccga
     4321 gatcggcccg cgcatggccg agttgagcgg ttcccggctg gccgcgcagc aacagatgga
     4381 aggcctcctg gcgccgcacc ggcccaagga gcccgcgtgg ttcctggcca ccgtcggcgt
     4441 ctcgcccgac caccagggca agggtctggg cagcgccgtc gtgctccccg gagtggaggc
     4501 ggccgagcgc gccggggtgc ccgccttcct ggagacctcc gcgccccgca acctcccctt
     4561 ctacgagcgg ctcggcttca ccgtcaccgc cgacgtcgag gtgcccgaag gaccgcgcac
     4621 ctggtgcatg acccgcaagc ccggtgccgg atcgggagag ggcagaggaa gtctgctaac
     4681 atgcggtgac gtcgaggaga atcctggccc accggtcgcc accagcgagc tgattaagga
     4741 gaacatgcac atgaagctgt acatggaggg caccgtggac aaccatcact tcaagtgcac
     4801 atccgagggc gaaggcaagc cctacgaggg cacccagacc atgagaatca aggtggtcga
     4861 gggcggccct ctccccttcg ccttcgacat cctggctact agcttcctct acggcagcaa
     4921 gaccttcatc aaccacaccc agggcatccc cgacttcttc aagcagtcct tccctgaggg
     4981 cttcacatgg gagagagtca ccacatacga agacgggggc gtgctgaccg ctacccagga
     5041 caccagcctc caggacggct gcctcatcta caacgtcaag atcagagggg tgaacttcac
     5101 atccaacggc cctgtgatgc agaagaaaac actcggctgg gaggccttca ccgagacgct
     5161 gtaccccgct gacggcggcc tggaaggcag aaacgacatg gccctgaagc tcgtgggcgg
     5221 gagccatctg atcgcaaaca tcaagaccac atatagatcc aagaaacccg ctaagaacct
     5281 caagatgcct ggcgtctact atgtggacta cagactggaa agaatcaagg aggccaacaa
     5341 cgagacctac gtcgagcagc acgaggtggc agtggccaga tactgcgacc tccctagcaa
     5401 actggggcac aagcttaatt aagaattcgt cgagggacct aataacttcg tatagcatac
     5461 attatacgaa gttatacatg tttaagggtt ccggttccac taggtacaat tcgatatcaa
     5521 gcttatcgat aatcaacctc tggattacaa aatttgtgaa agattgactg gtattcttaa
     5581 ctatgttgct ccttttacgc tatgtggata cgctgcttta atgcctttgt atcatgctat
     5641 tgcttcccgt atggctttca ttttctcctc cttgtataaa tcctggttgc tgtctcttta
     5701 tgaggagttg tggcccgttg tcaggcaacg tggcgtggtg tgcactgtgt ttgctgacgc
     5761 aacccccact ggttggggca ttgccaccac ctgtcagctc ctttccggga ctttcgcttt
     5821 ccccctccct attgccacgg cggaactcat cgccgcctgc cttgcccgct gctggacagg
     5881 ggctcggctg ttgggcactg acaattccgt ggtgttgtcg gggaaatcat cgtcctttcc
     5941 ttggctgctc gcctgtgttg ccacctggat tctgcgcggg acgtccttct gctacgtccc
     6001 ttcggccctc aatccagcgg accttccttc ccgcggcctg ctgccggctc tgcggcctct
     6061 tccgcgtctt cgccttcgcc ctcagacgag tcggatctcc ctttgggccg cctccccgca
     6121 tcgataccgt cgacctcgat cgagacctag aaaaacatgg agcaatcaca agtagcaata
     6181 cagcagctac caatgctgat tgtgcctggc tagaagcaca agaggaggag gaggtgggtt
     6241 ttccagtcac acctcaggta cctttaagac caatgactta caaggcagct gtagatctta
     6301 gccacttttt aaaagaaaag gggggactgg aagggctaat tcactcccaa cgaagacaag
     6361 atatccttga tctgtggatc taccacacac aaggctactt ccctgattgg cagaactaca
     6421 caccagggcc agggatcaga tatccactga cctttggatg gtgctacaag ctagtaccag
     6481 ttgagcaaga gaaggtagaa gaagccaatg aaggagagaa cacccgcttg ttacaccctg
     6541 tgagcctgca tgggatggat gacccggaga gagaagtatt agagtggagg tttgacagcc
     6601 gcctagcatt tcatcacatg gcccgagagc tgcatccgga ctgtactggg tctctctggt
     6661 tagaccagat ctgagcctgg gagctctctg gctaactagg gaacccactg cttaagcctc
     6721 aataaagctt gccttgagtg cttcaagtag tgtgtgcccg tctgttgtgt gactctggta
     6781 actagagatc cctcagaccc ttttagtcag tgtggaaaat ctctagcagc atgtgagcaa
     6841 aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc
     6901 tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga
     6961 caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc
     7021 cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt
     7081 ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct
     7141 gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg
     7201 agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta
     7261 gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct
     7321 acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa
     7381 gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt
     7441 gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta
     7501 cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat
     7561 caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa
     7621 gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct
     7681 cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta
     7741 cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct
     7801 caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg
     7861 gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa
     7921 gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt
     7981 cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta
     8041 catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca
     8101 gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta
     8161 ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct
     8221 gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg
     8281 cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac
     8341 tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact
     8401 gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa
     8461 atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt
     8521 ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat
     8581 gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg
     8641 acgtcgacgg atcgggagat ctcccgatcc cctatggtgc actctcagta caatctgctc
     8701 tgatgccgca tagttaagcc agtatctgct ccctgcttgt gtgttggagg tcgctgagta
     8761 gtgcgcgagc aaaatttaag ctacaacaag gcaaggcttg accgacaatt gcatgaagaa
     8821 tctgcttagg gttaggcgtt ttgcgctgct tcgcgatgta cgggccagat atacgcgtt
//
