LOCUS       Exported                8364 bp DNA     circular SYN 23-JUL-2020
DEFINITION  synthetic circular DNA
ACCESSION   urn.local...b-6pix0yf
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8364)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 14, 2021 from SnapGene 5.3.2
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..8364
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     rep_origin      complement(502..1117)
                     /direction=LEFT
                     /label=pUC Ori
     primer_bind     complement(1128..1140)
                     /label=zs-clon-047_gib-t2a-frag-fwd
     LTR             1465..1877
                     /label=5' LTR
     misc_feature    1928..2065
                     /label=psi
     primer_bind     1944..1961
                     /label=zs_seq_006
     misc_feature    2032..2396
                     /label=gag
     primer_bind     complement(2062..2082)
                     /label=zs_seq_005
     misc_feature    2542..2783
                     /label=RRE
     primer_bind     2547..2567
                     /label=zs_seq_007
     primer_bind     3148..3170
                     /label=zs_seq_008
     misc_feature    3184..3301
                     /label=cPPT
     misc_feature    3353..3354
                     /label=origin
     promoter        3360..4618
                     /label=EF-1a promoter
                     /label=EF-1a
     primer_bind     3420..3471
                     /label=zs-clon-013_ef1a-Fragment 1.FOR
     primer_bind     complement(3428..3478)
                     /label=zs-clon-012_ef1a-Vector.REV
     primer_bind     3550..3568
                     /label=zs_seq_001
     primer_bind     complement(3651..3670)
                     /label=zs-clon-028_cropseq-Vec.REVa
     primer_bind     complement(3651..3670)
                     /label=zs-clon-028_cropseq-Vec.REV
     primer_bind     complement(3812..3821)
                     /label=zs-clon-039_gib-N-Intein-Fragment.FOR
     primer_bind     4610..4629
                     /label=zs-clon-027_cropseq-Vec.FWDa
     primer_bind     4610..4629
                     /label=zs-clon-027_cropseq-Vec.FWD
     primer_bind     4625..4651
                     /label=zs-clon-023_gib-puro-Vector.FOR
     CDS             4631..5230
                     /label=Puro
     primer_bind     4634..4655
                     /label=zs-clon-011_t2a-puro-Vector.FOR
     primer_bind     5069..5086
                     /label=zs_seq_002
     primer_bind     complement(5115..5125)
                     /label=zs-clon-005_gib-mcherry-fwd
     primer_bind     5208..5227
                     /label=zs-clon-047_gib-t2a-frag-fwd
     primer_bind     complement(5213..5227)
                     /label=zs_clon_030
     primer_bind     5227..5249
                     /label=zs-clon-007_gib-tNGFR-Vector.FOR
     primer_bind     5228..5254
                     /label=zs-clon-003_gib-mch-vector-fwd
     primer_bind     5230..5254
                     /label=zs-clon-037_gib-N-Intein-CROPseq-Vector.FOR
     primer_bind     5230..5252
                     /label=zs-clon-029_gib-cropseq-backbone-Vector.FOR
     misc_RNA        5246..5834
                     /label=WPRE
     primer_bind     5606..5625
                     /label=gRNA validation primer 5'-TTGGGCACTGACAATTCCGT-3'
     primer_bind     5666..5683
                     /label=zs_seq_003
     LTR             5905..5940
                     /label=3' LTR
     promoter        5990..6234
                     /label=hU6 promoter
                     /label=hU6
     misc_feature    6200..6238
                     /label=39 bp gibson overlap
     primer_bind     6205..6239
                     /label=zs-clon-049_sgrna-cloning-mcsv-fwd
     misc_binding    6235..6238
                     /label=filler
                     /label=filler (1,885 bp)
     misc_feature    6260..6351
                     /label=gRNA backbone
     primer_bind     complement(6260..6307)
                     /label=zs-clon-050_sgrna-cloning-mcsv-rev
     misc_feature    6264
                     /label=A-U flip
     misc_feature    6272..6276
                     /label=stem extension part I
     misc_feature    6281..6285
                     /label=stem extension part II
     LTR             6380..6579
                     /label=3' LTR
     misc_feature    complement(6481..6501)
                     /label=MG348_sgOPTI_CROPsanger
     polyA_signal    6656..6786
                     /label=SV40 polyA
     primer_bind     complement(6845..6862)
                     /label=zs_seq_004
     CDS             join(7934..8364,1..376)
                     /label=AmpR
ORIGIN
        1 ctggatggag gcggataaag ttgcaggacc acttctgcgc tcggcccttc cggctggctg
       61 gtttattgct gataaatctg gagccggtga gcgtgggtct cgcggtatca ttgcagcact
      121 ggggccagat ggtaagccct cccgtatcgt agttatctac acgacgggga gtcaggcaac
      181 tatggatgaa cgaaatagac agatcgctga gataggtgcc tcactgatta agcattggta
      241 actgtcagac caagtttact catatatact ttagattgat ttaaaacttc atttttaatt
      301 taaaaggatc taggtgaaga tcctttttga taatctcatg accaaaatcc cttaacgtga
      361 gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc
      421 tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt
      481 ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc
      541 gcagatacca aatactgttc ttctagtgta gccgtagtta ggccaccact tcaagaactc
      601 tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg
      661 cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg
      721 gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga
      781 actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc
      841 ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg
      901 gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg
      961 atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt
     1021 tttacggttc ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc
     1081 tgattctgtg gataaccgta ttaccgcctt tgagtgagct gataccgctc gccgcagccg
     1141 aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa gagcgcccaa tacgcaaacc
     1201 gcctctcccc gcgcgttggc cgattcatta atgcagctgg cacgacaggt ttcccgactg
     1261 gaaagcgggc agtgagcgca acgcaattaa tgtgagttag ctcactcatt aggcacccca
     1321 ggctttacac tttatgcttc cggctcgtat gttgtgtgga attgtgagcg gataacaatt
     1381 tcacacagga aacagctatg accatgatta cgccaagcgc gcaattaacc ctcactaaag
     1441 ggaacaaaag ctggagctgc aagcttaatg tagtcttatg caatactctt gtagtcttgc
     1501 aacatggtaa cgatgagtta gcaacatgcc ttacaaggag agaaaaagca ccgtgcatgc
     1561 cgattggtgg aagtaaggtg gtacgatcgt gccttattag gaaggcaaca gacgggtctg
     1621 acatggattg gacgaaccac tgaattgccg cattgcagag atattgtatt taagtgccta
     1681 gctcgataca taaacgggtc tctctggtta gaccagatct gagcctggga gctctctggc
     1741 taactaggga acccactgct taagcctcaa taaagcttgc cttgagtgct tcaagtagtg
     1801 tgtgcccgtc tgttgtgtga ctctggtaac tagagatccc tcagaccctt ttagtcagtg
     1861 tggaaaatct ctagcagtgg cgcccgaaca gggacttgaa agcgaaaggg aaaccagagg
     1921 agctctctcg acgcaggact cggcttgctg aagcgcgcac ggcaagaggc gaggggcggc
     1981 gactggtgag tacgccaaaa attttgacta gcggaggcta gaaggagaga gatgggtgcg
     2041 agagcgtcag tattaagcgg gggagaatta gatcgcgatg ggaaaaaatt cggttaaggc
     2101 cagggggaaa gaaaaaatat aaattaaaac atatagtatg ggcaagcagg gagctagaac
     2161 gattcgcagt taatcctggc ctgttagaaa catcagaagg ctgtagacaa atactgggac
     2221 agctacaacc atcccttcag acaggatcag aagaacttag atcattatat aatacagtag
     2281 caaccctcta ttgtgtgcat caaaggatag agataaaaga caccaaggaa gctttagaca
     2341 agatagagga agagcaaaac aaaagtaaga ccaccgcaca gcaagcggcc gctgatcttc
     2401 agacctggag gaggagatat gagggacaat tggagaagtg aattatataa atataaagta
     2461 gtaaaaattg aaccattagg agtagcaccc accaaggcaa agagaagagt ggtgcagaga
     2521 gaaaaaagag cagtgggaat aggagctttg ttccttgggt tcttgggagc agcaggaagc
     2581 actatgggcg cagcgtcaat gacgctgacg gtacaggcca gacaattatt gtctggtata
     2641 gtgcagcagc agaacaattt gctgagggct attgaggcgc aacagcatct gttgcaactc
     2701 acagtctggg gcatcaagca gctccaggca agaatcctgg ctgtggaaag atacctaaag
     2761 gatcaacagc tcctggggat ttggggttgc tctggaaaac tcatttgcac cactgctgtg
     2821 ccttggaatg ctagttggag taataaatct ctggaacaga tttggaatca cacgacctgg
     2881 atggagtggg acagagaaat taacaattac acaagcttaa tacactcctt aattgaagaa
     2941 tcgcaaaacc agcaagaaaa gaatgaacaa gaattattgg aattagataa atgggcaagt
     3001 ttgtggaatt ggtttaacat aacaaattgg ctgtggtata taaaattatt cataatgata
     3061 gtaggaggct tggtaggttt aagaatagtt tttgctgtac tttctatagt gaatagagtt
     3121 aggcagggat attcaccatt atcgtttcag aatcttgaga caaatggcag tattcatcca
     3181 caattttaaa agaaaagggg ggattggggg gtacagtgca ggggaaagaa tagtagacat
     3241 aatagcaaca gacatacaaa ctaaagaatt acaaaaacaa attacaaaaa ttcaaaattt
     3301 tcgggtttat tacagggaca gcagagatcc actttggcgc cggctcgagg gggcccgggt
     3361 gcaaagatgg ataaagtttt aaacagagag gaatctttgc agctaatgga ccttctaggt
     3421 cttgaaagga gtgggaattg gctccggtgc ccgtcagtgg gcagagcgca catcgcccac
     3481 agtccccgag aagttggggg gaggggtcgg caattgatcc ggtgcctaga gaaggtggcg
     3541 cggggtaaac tgggaaagtg atgtcgtgta ctggctccgc ctttttcccg agggtggggg
     3601 agaaccgtat ataagtgcag tagtcgccgt gaacgttctt tttcgcaacg ggtttgccgc
     3661 cagaacacag gtaagtgccg tgtgtggttc ccgcgggcct ggcctcttta cgggttatgg
     3721 cccttgcgtg ccttgaatta cttccacctg gctgcagtac gtgattcttg atcccgagct
     3781 tcgggttgga agtgggtggg agagttcgag gccttgcgct taaggagccc cttcgcctcg
     3841 tgcttgagtt gaggcctggc ctgggcgctg gggccgccgc gtgcgaatct ggtggcacct
     3901 tcgcgcctgt ctcgctgctt tcgataagtc tctagccatt taaaattttt gatgacctgc
     3961 tgcgacgctt tttttctggc aagatagtct tgtaaatgcg ggccaagatc tgcacactgg
     4021 tatttcggtt tttggggccg cgggcggcga cggggcccgt gcgtcccagc gcacatgttc
     4081 ggcgaggcgg ggcctgcgag cgcggccacc gagaatcgga cgggggtagt ctcaagctgg
     4141 ccggcctgct ctggtgcctg gcctcgcgcc gccgtgtatc gccccgccct gggcggcaag
     4201 gctggcccgg tcggcaccag ttgcgtgagc ggaaagatgg ccgcttcccg gccctgctgc
     4261 agggagctca aaatggagga cgcggcgctc gggagagcgg gcgggtgagt cacccacaca
     4321 aaggaaaagg gcctttccgt cctcagccgt cgcttcatgt gactccacgg agtaccgggc
     4381 gccgtccagg cacctcgatt agttctcgag cttttggagt acgtcgtctt taggttgggg
     4441 ggaggggttt tatgcgatgg agtttcccca cactgagtgg gtggagactg aagttaggcc
     4501 agcttggcac ttgatgtaat tctccttgga atttgccctt tttgagtttg gatcttggtt
     4561 cattctcaag cctcagacag tggttcaaag tttttttctt ccatttcagg tgtcgtgacg
     4621 tacggccacc atgaccgagt acaagcccac ggtgcgcctc gccacccgcg acgacgtccc
     4681 cagggccgta cgcaccctcg ccgccgcgtt cgccgactac cccgccacgc gccacaccgt
     4741 cgatccggac cgccacatcg agcgggtcac cgagctgcaa gaactcttcc tcacgcgcgt
     4801 cgggctcgac atcggcaagg tgtgggtcgc ggacgacggc gccgccgtgg cggtctggac
     4861 cacgccggag agcgtcgaag cgggggcggt gttcgccgag atcggcccgc gcatggccga
     4921 gttgagcggt tcccggctgg ccgcgcagca acagatggaa ggcctcctgg cgccgcaccg
     4981 gcccaaggag cccgcgtggt tcctggccac cgtcggagtc tcgcccgacc accagggcaa
     5041 gggtctgggc agcgccgtcg tgctccccgg agtggaggcg gccgagcgcg ccggggtgcc
     5101 cgccttcctg gagacctccg cgccccgcaa cctccccttc tacgagcggc tcggcttcac
     5161 cgtcaccgcc gacgtcgagg tgcccgaagg accgcgcacc tggtgcatga cccgcaagcc
     5221 cggtgcctga acgcgttaag tcgacaatca acctctggat tacaaaattt gtgaaagatt
     5281 gactggtatt cttaactatg ttgctccttt tacgctatgt ggatacgctg ctttaatgcc
     5341 tttgtatcat gctattgctt cccgtatggc tttcattttc tcctccttgt ataaatcctg
     5401 gttgctgtct ctttatgagg agttgtggcc cgttgtcagg caacgtggcg tggtgtgcac
     5461 tgtgtttgct gacgcaaccc ccactggttg gggcattgcc accacctgtc agctcctttc
     5521 cgggactttc gctttccccc tccctattgc cacggcggaa ctcatcgccg cctgccttgc
     5581 ccgctgctgg acaggggctc ggctgttggg cactgacaat tccgtggtgt tgtcggggaa
     5641 atcatcgtcc tttccttggc tgctcgcctg tgttgccacc tggattctgc gcgggacgtc
     5701 cttctgctac gtcccttcgg ccctcaatcc agcggacctt ccttcccgcg gcctgctgcc
     5761 ggctctgcgg cctcttccgc gtcttcgcct tcgccctcag acgagtcgga tctccctttg
     5821 ggccgcctcc ccgcgtcgac tttaagacca atgacttaca aggcagctgt agatcttagc
     5881 cactttttaa aagaaaaggg gggactggaa gggctaattc actcccaacg aagacaagat
     5941 ttcaccatta tcgtttcaga cccacctccc aaccccgagg ggacccagag agggcctatt
     6001 tcccatgatt ccttcatatt tgcatatacg atacaaggct gttagagaga taattagaat
     6061 taatttgact gtaaacacaa agatattagt acaaaatacg tgacgtagaa agtaataatt
     6121 tcttgggtag tttgcagttt taaaattatg ttttaaaatg gactatcata tgcttaccgt
     6181 aacttgaaag tatttcgatt tcttggcttt atatatcttg tggaaaggac gaaacaccga
     6241 acaaagcacc ctccccactg tttaagagct atgctggaaa cagcatagca agtttaaata
     6301 aggctagtcc gttatcaact tgaaaaagtg gcaccgagtc ggtgcttttt taagcttggc
     6361 gtaactagat cttgagacac tgctttttgc ttgtactggg tctctctggt tagaccagat
     6421 ctgagcctgg gagctctctg gctaactagg gaacccactg cttaagcctc aataaagctt
     6481 gccttgagtg cttcaagtag tgtgtgcccg tctgttgtgt gactctggta actagagatc
     6541 cctcagaccc ttttagtcag tgtggaaaat ctctagcagt acgtatagta gttcatgtca
     6601 tcttattatt cagtatttat aacttgcaaa gaaatgaata tcagagagtg agaggaactt
     6661 gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt tcacaaataa
     6721 agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca
     6781 tgtctggctc tagctatccc gcccctaact ccgcccatcc cgcccctaac tccgcccagt
     6841 tccgcccatt ctccgcccca tggctgacta atttttttta tttatgcaga ggccgaggcc
     6901 gcctcggcct ctgagctatt ccagaagtag tgaggaggct tttttggagg cctagggacg
     6961 tacccaattc gccctatagt gagtcgtatt acgcgcgctc actggccgtc gttttacaac
     7021 gtcgtgactg ggaaaaccct ggcgttaccc aacttaatcg ccttgcagca catccccctt
     7081 tcgccagctg gcgtaatagc gaagaggccc gcaccgatcg cccttcccaa cagttgcgca
     7141 gcctgaatgg cgaatgggac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg
     7201 ttacgcgcag cgtgaccgct acacttgcca gcgccctagc gcccgctcct ttcgctttct
     7261 tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat cgggggctcc
     7321 ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt gattagggtg
     7381 atggttcacg tagtgggcca tcgccctgat agacggtttt tcgccctttg acgttggagt
     7441 ccacgttctt taatagtgga ctcttgttcc aaactggaac aacactcaac cctatctcgg
     7501 tctattcttt tgatttataa gggattttgc cgatttcggc ctattggtta aaaaatgagc
     7561 tgatttaaca aaaatttaac gcgaatttta acaaaatatt aacgcttaca atttaggtgg
     7621 cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa
     7681 tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa
     7741 gagtatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg cattttgcct
     7801 tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg
     7861 tgcacgagtg ggttacatcg aactggatct caacagcggt aagatccttg agagttttcg
     7921 ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt
     7981 atcccgtatt gacgccgggc aagagcaact cggtcgccgc atacactatt ctcagaatga
     8041 cttggttgag tactcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga
     8101 attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac
     8161 gatcggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc atgtaactcg
     8221 ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc gtgacaccac
     8281 gatgcctgta gcaatggcaa caacgttgcg caaactatta actggcgaac tacttactct
     8341 agcttcccgg caacaattaa taga
//
