LOCUS       Exported                8246 bp DNA     circular SYN 19-MAY-2022
DEFINITION  .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8246)
  AUTHORS   Frisen Lab
  TITLE     Direct Submission
  JOURNAL   Exported May 19, 2022 from SnapGene 6.0.2
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..8246
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     source          2485..2500
                     /mol_type="other DNA"
                     /note="color: #ffffff"
                     /organism="lentiGuide-Puro"
     source          2641..3862
                     /mol_type="other DNA"
                     /note="dnas_title=dCAS9-VP64_GFP"
                     /note="color: #ffffff"
                     /organism="dCAS9-VP64_GFP"
     source          5052..5067
                     /mol_type="other DNA"
                     /note="color: #ffffff"
                     /organism="lentiGuide-Puro"
     protein_bind    124..145
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
                     /note="color: #31849b"
     promoter        160..190
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
                     /note="This forward directional feature has 3 segments:
                        1: 160 .. 165 / #ffffff / -35
                        2: 166 .. 183 / #ffffff
                        3: 184 .. 190 / #ffffff / -10"
     protein_bind    198..214
                     /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)."
                     /note="color: #31849b"
     primer_bind     222..238
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
                     /note="color: #a020f0; direction: RIGHT"
     promoter        259..277
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
                     /note="color: #ffffff; direction: RIGHT"
     promoter        305..531
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
                     /note="color: #ffffff; direction: RIGHT"
     LTR             532..712
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
                     /note="color: #ffe4c4"
     misc_feature    755..800
                     /label=RNA packaging signal
                     /note="RNA packaging signal"
                     /note="color: #eaff50"
     misc_feature    759..884
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
                     /note="color: #ffe4c4"
     misc_feature    868..1232
                     /label=gag
                     /note="gag"
                     /note="color: #eaff50; direction: RIGHT"
     misc_feature    1377..1610
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
                     /note="color: #ffe4c4"
     promoter        2137..2377
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA"
                     /note="color: #ffffff; direction: RIGHT"
     misc_feature    2388..2435
                     /label=shScramble
                     /note="color: #a6acb3"
     misc_feature    2409..2414
                     /label=loop
                     /note="color: #a6acb3"
     misc_feature    2415..2435
                     /label=antisense
                     /note="color: #a6acb3"
     misc_feature    2481..2598
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
                     /note="color: #ffe4c4"
     misc_feature    2485..2500
                     /label=cPPT
                     /note="color: #a6acb3"
     promoter        2666..3844
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
                     /note="color: #ffffff; direction: RIGHT"
     intron          2897..3835
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
                     /note="color: #------"
     misc_feature    3851..3856
                     /label=Kozak
                     /note="color: #a6acb3"
     misc_feature    3857..3859
                     /label=start
                     /note="color: #a6acb3"
     CDS             3875..4251
                     /label=Histone H2b CDS
                     /note="Histone H2b CDS  (Kanda et al, Curr. Biol 8: 377-85;
                     Clone used is 98.4% identical to X00088)"
                     /note="color: #0000ff; direction: RIGHT"
     CDS             4271..4990
                     /label=EGFP
                     /note="EGFP"
                     /note="color: #00ff00; direction: RIGHT"
     misc_feature    4966..4995
                     /label=MRX1680_LHA
                     /note="color: #a6acb3"
     misc_feature    4996..5002
                     /label=future barcode
                     /note="color: #ff00ff"
     misc_feature    5003..5046
                     /label=MRX1680_RHA
                     /note="color: #a6acb3"
     misc_feature    5052..5067
                     /label=cPPT
                     /note="color: #a6acb3"
     LTR             5069..5302
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
                     /note="color: #ffe4c4"
     misc_feature    5195..5200
                     /label=PAS
                     /note="color: #a6acb3"
     misc_feature    5219..5248
                     /label=future polyA end
                     /note="color: #a6acb3"
     polyA_signal    5374..5495
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
                     /note="color: #a6acb3"
     rep_origin      5535..5670
                     /label=SV40 ori
                     /note="SV40 origin of replication"
                     /note="color: #ffff00"
     promoter        complement(5691..5709)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
                     /note="color: #ffffff; direction: LEFT"
     primer_bind     complement(5719..5735)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
                     /note="color: #a020f0; direction: LEFT"
     rep_origin      5877..6332
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
                     /note="color: #ffff00"
     promoter        6358..6462
                     /gene="bla"
                     /label=AmpR promoter
                     /note="color: #ffffff; direction: RIGHT"
     CDS             6463..7323
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /note="This feature has 2 segments:
                        1: 6463 .. 6531 / #ccffcc / signal sequence
                        2: 6532 .. 7323 / #ccffcc
                     Cleavage site after base 6531"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      7494..8082
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
                     /note="color: #ffff00"
ORIGIN
        1 gagcgaggaa gcggaagagc gcccaatacg caaaccgcct ctccccgcgc gttggccgat
       61 tcattaatgc agctggcacg acaggtttcc cgactggaaa gcgggcagtg agcgcaacgc
      121 aattaatgtg agttagctca ctcattaggc accccaggct ttacacttta tgcttccggc
      181 tcgtatgttg tgtggaattg tgagcggata acaatttcac acaggaaaca gctatgacca
      241 tgattacgcc aagcgcgcaa ttaaccctca ctaaagggaa caaaagctgg agctgcaagc
      301 ttaatgtagt cttatgcaat actcttgtag tcttgcaaca tggtaacgat gagttagcaa
      361 catgccttac aaggagagaa aaagcaccgt gcatgccgat tggtggaagt aaggtggtac
      421 gatcgtgcct tattaggaag gcaacagacg ggtctgacat ggattggacg aaccactgaa
      481 ttgccgcatt gcagagatat tgtatttaag tgcctagctc gatacataaa cgggtctctc
      541 tggttagacc agatctgagc ctgggagctc tctggctaac tagggaaccc actgcttaag
      601 cctcaataaa gcttgccttg agtgcttcaa gtagtgtgtg cccgtctgtt gtgtgactct
      661 ggtaactaga gatccctcag acccttttag tcagtgtgga aaatctctag cagtggcgcc
      721 cgaacaggga cttgaaagcg aaagggaaac cagaggagct ctctcgacgc aggactcggc
      781 ttgctgaagc gcgcacggca agaggcgagg ggcggcgact ggtgagtacg ccaaaaattt
      841 tgactagcgg aggctagaag gagagagatg ggtgcgagag cgtcagtatt aagcggggga
      901 gaattagatc gcgatgggaa aaaattcggt taaggccagg gggaaagaaa aaatataaat
      961 taaaacatat agtatgggca agcagggagc tagaacgatt cgcagttaat cctggcctgt
     1021 tagaaacatc agaaggctgt agacaaatac tgggacagct acaaccatcc cttcagacag
     1081 gatcagaaga acttagatca ttatataata cagtagcaac cctctattgt gtgcatcaaa
     1141 ggatagagat aaaagacacc aaggaagctt tagacaagat agaggaagag caaaacaaaa
     1201 gtaagaccac cgcacagcaa gcggccgctg atcttcagac ctggaggagg agatatgagg
     1261 gacaattgga gaagtgaatt atataaatat aaagtagtaa aaattgaacc attaggagta
     1321 gcacccacca aggcaaagag aagagtggtg cagagagaaa aaagagcagt gggaatagga
     1381 gctttgttcc ttgggttctt gggagcagca ggaagcacta tgggcgcagc gtcaatgacg
     1441 ctgacggtac aggccagaca attattgtct ggtatagtgc agcagcagaa caatttgctg
     1501 agggctattg aggcgcaaca gcatctgttg caactcacag tctggggcat caagcagctc
     1561 caggcaagaa tcctggctgt ggaaagatac ctaaaggatc aacagctcct ggggatttgg
     1621 ggttgctctg gaaaactcat ttgcaccact gctgtgcctt ggaatgctag ttggagtaat
     1681 aaatctctgg aacagatttg gaatcacacg acctggatgg agtgggacag agaaattaac
     1741 aattacacaa gcttaataca ctccttaatt gaagaatcgc aaaaccagca agaaaagaat
     1801 gaacaagaat tattggaatt agataaatgg gcaagtttgt ggaattggtt taacataaca
     1861 aattggctgt ggtatataaa attattcata atgatagtag gaggcttggt aggtttaaga
     1921 atagtttttg ctgtactttc tatagtgaat agagttaggc agggatattc accattatcg
     1981 tttcagaccc acctcccaac cccgagggga cccgacaggc ccgaaggaat agaagaagaa
     2041 ggtggagaga gagacagaga cagatccatt cgattagtga acggatctcg acggtatcga
     2101 tcacgagact agcctcgagc ggccgccccc ttcaccgagg gcctatttcc catgattcct
     2161 tcatatttgc atatacgata caaggctgtt agagagataa ttggaattaa tttgactgta
     2221 aacacaaaga tattagtaca aaatacgtga cgtagaaagt aataatttct tgggtagttt
     2281 gcagttttaa aattatgttt taaaatggac tatcatatgc ttaccgtaac ttgaaagtat
     2341 ttcgatttct tggctttata tatcttgtgg aaaggacgaa acaccggcaa caagatgaag
     2401 agcaccaact cgagttggtg ctcttcatct tgttgttttt aattctcgac ctcgagacaa
     2461 atggcagtat tcatccacaa ttttaaaaga aaagggggga ttggggggta cagtgcaggg
     2521 gaaagaatag tagacataat agcaacagac atacaaacta aagaattaca aaaacaaatt
     2581 acaaaaattc aaaattttcg ggtttattac agggacagca gagatccact ttggactagt
     2641 ctaggtcttg aaaggagtgg gaattggctc cggtgcccgt cagtgggcag agcgcacatc
     2701 gcccacagtc cccgagaagt tggggggagg ggtcggcaat tgaaccggtg cctagagaag
     2761 gtggcgcggg gtaaactggg aaagtgatgt cgtgtactgg ctccgccttt ttcccgaggg
     2821 tgggggagaa ccgtatataa gtgcagtagt cgccgtgaac gttctttttc gcaacgggtt
     2881 tgccgccaga acacaggtaa gtgccgtgtg tggttcccgc gggcctggcc tctttacggg
     2941 ttatggccct tgcgtgcctt gaattacttc cacctggctg cagtacgtga ttcttgatcc
     3001 cgagcttcgg gttggaagtg ggtgggagag ttcgaggcct tgcgcttaag gagccccttc
     3061 gcctcgtgct tgagttgagg cctggcctgg gcgctggggc cgccgcgtgc gaatctggtg
     3121 gcaccttcgc gcctgtctcg ctgctttcga taagtctcta gccatttaaa atttttgatg
     3181 acctgctgcg acgctttttt tctggcaaga tagtcttgta aatgcgggcc aagatctgca
     3241 cactggtatt tcggtttttg gggccgcggg cggcgacggg gcccgtgcgt cccagcgcac
     3301 atgttcggcg aggcggggcc tgcgagcgcg gccaccgaga atcggacggg ggtagtctca
     3361 agctggccgg cctgctctgg tgcctggcct cgcgccgccg tgtatcgccc cgccctgggc
     3421 ggcaaggctg gcccggtcgg caccagttgc gtgagcggaa agatggccgc ttcccggccc
     3481 tgctgcaggg agctcaaaat ggaggacgcg gcgctcggga gagcgggcgg gtgagtcacc
     3541 cacacaaagg aaaagggcct ttccgtcctc agccgtcgct tcatgtgact ccacggagta
     3601 ccgggcgccg tccaggcacc tcgattagtt ctcgagcttt tggagtacgt cgtctttagg
     3661 ttggggggag gggttttatg cgatggagtt tccccacact gagtgggtgg agactgaagt
     3721 taggccagct tggcacttga tgtaattctc cttggaattt gccctttttg agtttggatc
     3781 ttggttcatt ctcaagcctc agacagtggt tcaaagtttt tttcttccat ttcaggtgtc
     3841 gtgacgtacg gccaccatga gcgtcgacgc caccatgcca gagccagcga agtctgctcc
     3901 cgccccgaaa aagggctcca agaaggcggt gactaaggcg cagaagaaag gcggcaagaa
     3961 gcgcaagcgc agccgcaagg agagctattc catctatgtg tacaaggttc tgaagcaggt
     4021 ccaccctgac accggcattt cgtccaaggc catgggcatc atgaactcgt ttgtgaacga
     4081 cattttcgag cgcatcgcag gtgaggcttc ccgcctggcg cattacaaca agcgctcgac
     4141 catcacctcc agggagatcc agacggccgt gcgcctgctg ctgcctgggg agttggccaa
     4201 gcacgccgtg tccgagggta ctaaggccat caccaagtac accagcgcta aggatccacc
     4261 ggtcgccacc atggtgagca agggcgagga gctgttcacc ggggtggtgc ccatcctggt
     4321 cgagctggac ggcgacgtaa acggccacaa gttcagcgtg tccggcgagg gcgagggcga
     4381 tgccacctac ggcaagctga ccctgaagtt catctgcacc accggcaagc tgcccgtgcc
     4441 ctggcccacc ctcgtgacca ccctgaccta cggcgtgcag tgcttcagcc gctaccccga
     4501 ccacatgaag cagcacgact tcttcaagtc cgccatgccc gaaggctacg tccaggagcg
     4561 caccatcttc ttcaaggacg acggcaacta caagacccgc gccgaggtga agttcgaggg
     4621 cgacaccctg gtgaaccgca tcgagctgaa gggcatcgac ttcaaggagg acggcaacat
     4681 cctggggcac aagctggagt acaactacaa cagccacaac gtctatatca tggccgacaa
     4741 gcagaagaac ggcatcaagg tgaacttcaa gatccgccac aacatcgagg acggcagcgt
     4801 gcagctcgcc gaccactacc agcagaacac ccccatcggc gacggccccg tgctgctgcc
     4861 cgacaaccac tacctgagca cccagtccgc cctgagcaaa gaccccaacg agaagcgcga
     4921 tcacatggtc ctgctggagt tcgtgaccgc cgccgggatc actctcggca tggacgagct
     4981 gtacaagtaa atgcatcggt acctttaaga ccaatgactt acaaggcagc tgtagatctt
     5041 agccactttt taaaagaaaa ggggggactg gaagggctaa ttcactccca acgaagacaa
     5101 gatctgcttt ttgcttgtac tgggtctctc tggttagacc agatctgagc ctgggagctc
     5161 tctggctaac tagggaaccc actgcttaag cctcaataaa gcttgccttg agtgcttcaa
     5221 gtagtgtgtg cccgtctgtt gtgtgactct ggtaactaga gatccctcag acccttttag
     5281 tcagtgtgga aaatctctag cagtagtagt tcatgtcatc ttattattca gtatttataa
     5341 cttgcaaaga aatgaatatc agagagtgag aggaacttgt ttattgcagc ttataatggt
     5401 tacaaataaa gcaatagcat cacaaatttc acaaataaag catttttttc actgcattct
     5461 agttgtggtt tgtccaaact catcaatgta tcttatcatg tctggctcta gctatcccgc
     5521 ccctaactcc gcccatcccg cccctaactc cgcccagttc cgcccattct ccgccccatg
     5581 gctgactaat tttttttatt tatgcagagg ccgaggccgc ctcggcctct gagctattcc
     5641 agaagtagtg aggaggcttt tttggaggcc tagggacgta cccaattcgc cctatagtga
     5701 gtcgtattac gcgcgctcac tggccgtcgt tttacaacgt cgtgactggg aaaaccctgg
     5761 cgttacccaa cttaatcgcc ttgcagcaca tccccctttc gccagctggc gtaatagcga
     5821 agaggcccgc accgatcgcc cttcccaaca gttgcgcagc ctgaatggcg aatgggacgc
     5881 gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac
     5941 acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt
     6001 cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc
     6061 tttacggcac ctcgacccca aaaaacttga ttagggtgat ggttcacgta gtgggccatc
     6121 gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact
     6181 cttgttccaa actggaacaa cactcaaccc tatctcggtc tattcttttg atttataagg
     6241 gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc
     6301 gaattttaac aaaatattaa cgcttacaat ttaggtggca cttttcgggg aaatgtgcgc
     6361 ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa
     6421 taaccctgat aaatgcttca ataatattga aaaaggaaga gtatgagtat tcaacatttc
     6481 cgtgtcgccc ttattccctt ttttgcggca ttttgccttc ctgtttttgc tcacccagaa
     6541 acgctggtga aagtaaaaga tgctgaagat cagttgggtg cacgagtggg ttacatcgaa
     6601 ctggatctca acagcggtaa gatccttgag agttttcgcc ccgaagaacg ttttccaatg
     6661 atgagcactt ttaaagttct gctatgtggc gcggtattat cccgtattga cgccgggcaa
     6721 gagcaactcg gtcgccgcat acactattct cagaatgact tggttgagta ctcaccagtc
     6781 acagaaaagc atcttacgga tggcatgaca gtaagagaat tatgcagtgc tgccataacc
     6841 atgagtgata acactgcggc caacttactt ctgacaacga tcggaggacc gaaggagcta
     6901 accgcttttt tgcacaacat gggggatcat gtaactcgcc ttgatcgttg ggaaccggag
     6961 ctgaatgaag ccataccaaa cgacgagcgt gacaccacga tgcctgtagc aatggcaaca
     7021 acgttgcgca aactattaac tggcgaacta cttactctag cttcccggca acaattaata
     7081 gactggatgg aggcggataa agttgcagga ccacttctgc gctcggccct tccggctggc
     7141 tggtttattg ctgataaatc tggagccggt gagcgtgggt ctcgcggtat cattgcagca
     7201 ctggggccag atggtaagcc ctcccgtatc gtagttatct acacgacggg gagtcaggca
     7261 actatggatg aacgaaatag acagatcgct gagataggtg cctcactgat taagcattgg
     7321 taactgtcag accaagttta ctcatatata ctttagattg atttaaaact tcatttttaa
     7381 tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt
     7441 gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat
     7501 cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg
     7561 gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga
     7621 gcgcagatac caaatactgt tcttctagtg tagccgtagt taggccacca cttcaagaac
     7681 tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt
     7741 ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag
     7801 cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc
     7861 gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag
     7921 gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca
     7981 gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt
     8041 cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc
     8101 tttttacggt tcctggcctt ttgctggcct tttgctcaca tgttctttcc tgcgttatcc
     8161 cctgattctg tggataaccg tattaccgcc tttgagtgag ctgataccgc tcgccgcagc
     8221 cgaacgaccg agcgcagcga gtcagt
//
