LOCUS       Exported                6945 bp ds-DNA     circular SYN 29-JUN-2021
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6945)
  AUTHORS   Trial User
  TITLE     Direct Submission
  JOURNAL   Exported Tuesday, Jun 29, 2021 from SnapGene 5.1.7
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6945
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     primer_bind     complement(229..248)
                     /label=AKH5032 pc
     primer_bind     1901..1957
                     /label=EAT327
     primer_bind     complement(1901..1957)
                     /label=EAT332
     primer_bind     2018..2045
                     /label=KAS017
     primer_bind     2050..2072
                     /label=ABR78
                     /note="hTert LTR rev"
     primer_bind     2637..2659
                     /label=LR31
     primer_bind     2637..2653
                     /label=KAS152
     misc_feature    complement(2641..2675)
                     /label=5' TR
     primer_bind     2641..2661
                     /label=KAS055
     primer_bind     2641..2659
                     /label=EAT487
     primer_bind     2678..2697
                     /label=KAS223
     primer_bind     2717..2768
                     /label=KAS039
     primer_bind     complement(2717..2768)
                     /label=KAS040
     primer_bind     complement(2720..2740)
                     /label=KAS041
     primer_bind     2720..2740
                     /label=KAS042
     primer_bind     2746..2765
                     /label=KAS043
     primer_bind     2921..2976
                     /label=Fragment 1.FOR
     primer_bind     2958..2977
                     /label=JW174
                     /note="JW174 pegRNA-PB-multi_FOR"
     primer_bind     2958..2977
                     /label=JW175
                     /note="JW175 pegRNA-PB-multi_REV"
     primer_bind     2960..2976
                     /label=JW13 (U6_promoter.FOR)
     promoter        2961..3205
                     /label=Human U6 Promoter
     promoter        2961..3201
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA"
     primer_bind     2961..2980
                     /label=EAT24
     primer_bind     2970..2989
                     /label=EAT58
     primer_bind     2990..3009
                     /label=EAT59
     primer_bind     3119..3161
                     /label=EAT20
     primer_bind     3128..3168
                     /label=EAT21
     primer_bind     3181..3205
                     /label=JW67 (hu6-filler FOR)
     primer_bind     3184..3226
                     /label=JW34 (peg accept. F)
     promoter        3184..3209
                     /standard_name="hU6"
                     /label=hU6 promoter
     primer_bind     3185..3209
                     /label=JW62 pegRNA-target F
     primer_bind     3190..3209
                     /label=EAT 708
                     /note="Library_NGS_Fwd"
     primer_bind     3190..3209
                     /label=EAT616
     primer_bind     3190..3209
                     /label=EAT56
     terminator      3217..3252
                     /standard_name="bidirectional terminator, weaker in 
                     forwarddirection"
     terminator      3219..3250
                     /label=tonB terminator
                     /note="bidirectional E. coli tonB-P14 transcription 
                     terminator"
     CDS             complement(3267..3944)
                     /standard_name="mRFP"
                     /label=mRFP
     misc_feature    complement(3945..3969)
                     /standard_name="RBS"
                     /label=RBS
     misc_feature    complement(3970..4023)
                     /standard_name="pTet"
                     /label=pTet
     protein_bind    3980..3998
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    4005..4023
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     complement(4008..4046)
                     /label=JW35 (peg accept. R)
     primer_bind     complement(4014..4037)
                     /label=JW139 pegRNA-GG_R
     primer_bind     4024..4055
                     /label=JW36 (cPPT -pT  F)
     primer_bind     complement(4027..4070)
                     /label=Fragment 1.REV
     primer_bind     complement(4028..4070)
                     /label=JW176
                     /note="JW176 pegRNA-GG_(PB)_REV"
     terminator      4031..4037
                     /standard_name="U6 terminator"
                     /label=U6 terminator
     primer_bind     4046..4080
                     /label=Fragment 2.FOR
     primer_bind     complement(4061..4084)
                     /label=JW37 (cPPT R)
     promoter        4064..5321
                     /label=EF-1a
     primer_bind     4071..4088
                     /label=LR298
     primer_bind     complement(4094..4116)
                     /label=EAT 709
                     /note="Library_NGS_Rev"
     primer_bind     complement(4094..4116)
                     /label=EAT617
     primer_bind     4119..4144
                     /label=EAT601
     primer_bind     4119..4139
                     /label=KAS218
     primer_bind     4120..4138
                     /label=EAT69
     primer_bind     complement(4124..4143)
                     /label=EAT618
     primer_bind     complement(4136..4155)
                     /label=EAT619
     primer_bind     4143..4161
                     /label=KAS329
                     /note="EF1a NEB fwd"
     primer_bind     4163..4180
                     /label=ABR91
                     /note="hTert neb fwd"
     primer_bind     complement(4166..4185)
                     /label=EAT23
     primer_bind     4218..4237
                     /label=KAS224
     primer_bind     complement(4240..4259)
                     /label=KAS333
                     /note="EF1-alpha (AS)"
     primer_bind     4314..4333
                     /label=EAT452
     primer_bind     4322..4341
                     /label=KAS478
                     /note="PB mapping EFS (S)"
     primer_bind     4348..4367
                     /label=MR3
     primer_bind     4352..4371
                     /label=KAS479
                     /note="PB mapping EFS nested (S)"
     primer_bind     complement(4356..4375)
                     /label=EAT602
     intron          4375..5312
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     5093..5114
                     /label=LR129
     primer_bind     5162..5181
                     /label=KAS334
                     /note="EF1-alpha (S)"
     primer_bind     complement(5293..5326)
                     /label=KAS330
                     /note="EF1a NEB rev"
     primer_bind     complement(5293..5321)
                     /label=KAS344
                     /note="EF1-a NEB 4 (AS)"
     primer_bind     complement(5298..5321)
                     /label=KAS345
                     /note="EF1-a NEB 5 (AS)"
     primer_bind     complement(5301..5330)
                     /label=AKH5189
     primer_bind     complement(5301..5321)
                     /label=AKH5185
     primer_bind     complement(5304..5326)
                     /label=KAS342
                     /note="EF1-a NEB (AS)"
     primer_bind     complement(5304..5326)
                     /label=KAS343
                     /note="EF1-a NEB 3 (AS)"
     primer_bind     complement(5304..5326)
                     /label=KAS341
                     /note="EF1-a NEB (AS)"
     CDS             5334..5933
                     /label=Puromycin resistance
     primer_bind     complement(5384..5403)
                     /label=AKH5022 pc
     primer_bind     complement(5414..5433)
                     /label=AKH5023 pc
     primer_bind     complement(5437..5456)
                     /label=KAS225
     primer_bind     complement(5450..5493)
                     /label=KAS271
     primer_bind     complement(5450..5479)
                     /label=KAS273
     primer_bind     5458..5485
                     /label=KAS272
     primer_bind     5462..5485
                     /label=KAS268
     primer_bind     complement(5671..5701)
                     /label=KAS249
     primer_bind     5676..5706
                     /label=KAS248
     primer_bind     complement(5710..5730)
                     /label=KAS145
     primer_bind     5710..5730
                     /label=KAS146
     primer_bind     5908..5927
                     /label=KAS318
                     /note="3'Puro (S)"
     primer_bind     complement(5912..5930)
                     /label=EAT352
     primer_bind     complement(5913..5930)
                     /label=KAS222
     primer_bind     complement(5914..5933)
                     /label=MNP104
     primer_bind     complement(5916..5935)
                     /label=EAT74
     primer_bind     complement(5916..5933)
                     /label=ABR95
                     /note="IL4 neb rev"
     primer_bind     complement(5916..5933)
                     /label=KAS283
     primer_bind     complement(5918..5964)
                     /label=KAS609
                     /note="Puro NEB (AS)"
     primer_bind     complement(5919..5934)
                     /label=JW133
                     /note="JW133 mCherry-IRES-Puro_R"
     primer_bind     complement(5923..5970)
                     /label=Fragment 2.REV
     primer_bind     6112..6129
                     /label=LR43
     primer_bind     6127..6146
                     /label=KAS057
     misc_feature    complement(6138..6200)
                     /label=3' TR
     primer_bind     complement(6177..6200)
                     /label=EAT486
     primer_bind     complement(6183..6205)
                     /label=KAS151
     primer_bind     complement(6185..6204)
                     /label=KAS83
     primer_bind     complement(6188..6204)
                     /label=KAS152
     primer_bind     complement(6370..6389)
                     /label=AKH5168
     primer_bind     complement(6405..6425)
                     /label=LR76
     primer_bind     complement(6729..6748)
                     /label=KAS143
ORIGIN
        1 gtggcacttt tcggggaaat gtgcgcggaa cccctatttg tttatttttc taaatacatt
       61 caaatatgta tccgctcatg agacaataac cctgataaat gcttcaataa tattgaaaaa
      121 ggaagagtat gagtattcaa catttccgtg tcgcccttat tccctttttt gcggcatttt
      181 gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt
      241 tgggtgcacg agtgggttac atcgaactgg atctcaacag cggtaagatc cttgagagtt
      301 ttcgccccga agaacgtttt ccaatgatga gcacttttaa agttctgcta tgtggcgcgg
      361 tattatcccg tattgacgcc gggcaagagc aactcggtcg ccgcatacac tattctcaga
      421 atgacttggt tgagtactca ccagtcacag aaaagcatct tacggatggc atgacagtaa
      481 gagaattatg cagtgctgcc ataaccatga gtgataacac tgcggccaac ttacttctga
      541 caacgatcgg aggaccgaag gagctaaccg cttttttgca caacatgggg gatcatgtaa
      601 ctcgccttga tcgttgggaa ccggagctga atgaagccat accaaacgac gagcgtgaca
      661 ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact attaactggc gaactactta
      721 ctctagcttc ccggcaacaa ttaatagact ggatggaggc ggataaagtt gcaggaccac
      781 ttctgcgctc ggcccttccg gctggctggt ttattgctga taaatctgga gccggtgagc
      841 gtgggtctcg cggtatcatt gcagcactgg ggccagatgg taagccctcc cgtatcgtag
      901 ttatctacac gacggggagt caggcaacta tggatgaacg aaatagacag atcgctgaga
      961 taggtgcctc actgattaag cattggtaac tgtcagacca agtttactca tatatacttt
     1021 agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc ctttttgata
     1081 atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag
     1141 aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa
     1201 caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt
     1261 ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgtcctt ctagtgtagc
     1321 cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa
     1381 tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa
     1441 gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc
     1501 ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa
     1561 gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa
     1621 caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg
     1681 ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc
     1741 tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg
     1801 ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg
     1861 agtgagctga taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg
     1921 aagcggaaga gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat
     1981 gcagctggca cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaatg
     2041 tgagttagct cactcattag gcaccccagg ctttacactt tatgcttccg gctcgtatgt
     2101 tgtgtggaat tgtgagcgga taacaatttc acacaggaaa cagctatgac catgattacg
     2161 ccaagcgcgc ccgccgggta actcacgggg tatccatgtc catttctgcg gcatccagcc
     2221 aggatacccg tcctcgctga cgtaatatcc cagcgccgca ccgctgtcat taatctgcac
     2281 accggcacgg cagttccggc tgtcgccggt attgttcggg ttgctgatgc gcttcgggct
     2341 gaccatccgg aactgtgtcc ggaaaagccg cgacgaactg gtatcccagg tggcctgaac
     2401 gaacagttca ccgttaaagg cgtgcatggc cacaccttcc cgaatcatca tggtaaacgt
     2461 gcgttttcgc tcaacgtcaa tgcagcagca gtcatcctcg gcaaactctt tccatgccgc
     2521 ttcaacctcg cgggaaaagg cacgggcttc ttcctccccg atgcccagat agcgccagct
     2581 tgggcgatga ctgagccgga aaaaagaccc gacgatatga tcctgatgca gctagattaa
     2641 ccctagaaag atagtctgcg taaaattgac gcatgcattc ttgaaatatt gctctctctt
     2701 tctaaatagc gcgaatccgt cgctgtgcat ttaggacatc tcagtcgccg cttggagctc
     2761 ccgtgaggcg tgcttgtcaa tgcggtaagt gtcactgatt ttgaactata acgaccgcgt
     2821 gagtcaaaat gacgcatgat tatcttttac gtgactttta agatttaact catacgataa
     2881 ttatattgtt atttcatgtt ctacttacgt gataacttat tatatatata ttttcttgtt
     2941 atagatatca agcttccaga gagggcctat ttcccatgat tccttcatat ttgcatatac
     3001 gatacaaggc tgttagagag ataattagaa ttaatttgac tgtaaacaca aagatattag
     3061 tacaaaatac gtgacgtaga aagtaataat ttcttgggta gtttgcagtt ttaaaattat
     3121 gttttaaaat ggactatcat atgcttaccg taacttgaaa gtatttcgat ttcttggctt
     3181 tatatatctt gtggaaagga cgaaacaccc gagacggtag tcaaaagcct ccggtcggag
     3241 gcttttgact tggctgagga gtgccattaa gcaccggtgg agtgacgacc ttcagcacgt
     3301 tcgtactgtt caacgatggt gtagtcttcg ttgtgggagg tgatgtccag tttgatgtcg
     3361 gttttgtaag cacccggcag ctgaaccggt tttttagcca tgtaggtggt tttaacttca
     3421 gcgtcgtagt gaccaccgtc tttcagtttc agacgcattt tgatttcacc tttcagagca
     3481 ccgtcttccg ggtacatacg ttcggtggaa gcttcccaac ccatggtttt tttctgcata
     3541 accggaccgt cggacgggaa gttggtacca cgcagtttaa ctttgtagat gaactcaccg
     3601 tcttgcaggg aggagtcctg ggtaacggta acaacaccac cgtcttcgaa gttcataaca
     3661 cgttcccatt tgaaaccttc cgggaaggac agtttcaggt agtccgggat gtcagccggg
     3721 tgtttaacgt aagctttgga accgtactgg aactgcgggg acaggatgtc ccaagcgaac
     3781 ggcagcggac cacctttggt aactttcagt ttagcggtct gggtaccttc gtacggacga
     3841 ccttcacctt caccttcgat ttcgaactcg tgaccgttaa cggaaccttc catacgaact
     3901 ttgaaacgca tgaactcttt gataacgtct tcgctactcg ccatagatcc tttctcctct
     3961 ttcagatccg tgctcagtat ctctatcact gatagggatg tcaatctcta tcactgatag
     4021 ggacgtctcg tttttttaag cttggcgtaa ctagatcccc gggtgcaaag atggataaag
     4081 ttttaaacag agaggaatct ttgcagctaa tggaccttct aggtcttgaa aggagtggga
     4141 attggctccg gtgcccgtca gtgggcagag cgcacatcgc ccacagtccc cgagaagttg
     4201 gggggagggg tcggcaattg atccggtgcc tagagaaggt ggcgcggggt aaactgggaa
     4261 agtgatgtcg tgtactggct ccgccttttt cccgagggtg ggggagaacc gtatataagt
     4321 gcagtagtcg ccgtgaacgt tctttttcgc aacgggtttg ccgccagaac acaggtaagt
     4381 gccgtgtgtg gttcccgcgg gcctggcctc tttacgggtt atggcccttg cgtgccttga
     4441 attacttcca ctggctgcag tacgtgattc ttgatcccga gcttcgggtt ggaagtgggt
     4501 gggagagttc gaggccttgc gcttaaggag ccccttcgcc tcgtgcttga gttgaggcct
     4561 ggcctgggcg ctggggccgc cgcgtgcgaa tctggtggca ccttcgcgcc tgtctcgctg
     4621 ctttcgataa gtctctagcc atttaaaatt tttgatgacc tgctgcgacg ctttttttct
     4681 ggcaagatag tcttgtaaat gcgggccaag atctgcacac tggtatttcg gtttttgggg
     4741 ccgcgggcgg cgacggggcc cgtgcgtccc agcgcacatg ttcggcgagg cggggcctgc
     4801 gagcgcggcc accgagaatc ggacgggggt agtctcaagc tggccggcct gctctggtgc
     4861 ctggcctcgc gccgccgtgt atcgccccgc cctgggcggc aaggctggcc cggtcggcac
     4921 cagttgcgtg agcggaaaga tggccgcttc ccggccctgc tgcagggagc tcaaaatgga
     4981 ggacgcggcg ctcgggagag cgggcgggtg agtcacccac acaaaggaaa agggcctttc
     5041 cgtcctcagc cgtcgcttca tgtgactcca cggagtaccg ggcgccgtcc aggcacctcg
     5101 attagttctc gagcttttgg agtacgtcgt ctttaggttg gggggagggg ttttatgcga
     5161 tggagtttcc ccacactgag tgggtggaga ctgaagttag gccagcttgg cacttgatgt
     5221 aattctcctt ggaatttgcc ctttttgagt ttggatcttg gttcattctc aagcctcaga
     5281 cagtggttca aagttttttt cttccatttc aggtgtcgtg acgtacggcc accatgaccg
     5341 agtacaagcc cacggtgcgc ctcgccaccc gcgacgacgt ccccagggcc gtacgcaccc
     5401 tcgccgccgc gttcgccgac taccccgcca cgcgccacac cgtcgatccg gaccgccaca
     5461 tcgagcgggt caccgagctg caagaactct tcctcacgcg cgtcgggctc gacatcggca
     5521 aggtgtgggt cgcggacgac ggcgccgccg tggcggtctg gaccacgccg gagagcgtcg
     5581 aagcgggggc ggtgttcgcc gagatcggcc cgcgcatggc cgagttgagc ggttcccggc
     5641 tggccgcgca gcaacagatg gaaggcctcc tggcgccgca ccggcccaag gagcccgcgt
     5701 ggttcctggc caccgtcgga gtctcgcccg accaccaggg caagggtctg ggcagcgccg
     5761 tcgtgctccc cggagtggag gcggccgagc gcgccggggt gcccgccttc ctggagacct
     5821 ccgcgccccg caacctcccc ttctacgagc ggctcggctt caccgtcacc gccgacgtcg
     5881 aggtgcccga aggaccgcgc acctggtgca tgacccgcaa gcccggtgcc tgaacgcgtt
     5941 aacccgggct gcaggaattc gataaaagtt ttgttacttt atagaagaaa ttttgagttt
     6001 ttgttttttt ttaataaata aataaacata aataaattgt ttgttgaatt tattattagt
     6061 atgtaagtgt aaatataata aaacttaata tctattcaaa ttaataaata aacctcgata
     6121 tacagaccga taaaacacat gcgtcaattt tacacatgat tatctttaac gtacgtcaca
     6181 atatgattat ctttctaggg ttaatctagc tgcgtgttct gcagcgtgtc gagcatcttc
     6241 atctgctcca tcacgctgta aaacacattt gcaccgcgag tctgcccgtc ctccacgggt
     6301 tcaaaaacgt gaatgaacga ggcgcgctca ctggccgtcg ttttacaacg tcgtgactgg
     6361 gaaaaccctg gcgttaccca acttaatcgc cttgcagcac atcccccttt cgccagctgg
     6421 cgtaatagcg aagaggcccg caccgatcgc ccttcccaac agttgcgcag cctgaatggc
     6481 gaatgggacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc
     6541 gtgaccgcta cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt
     6601 ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc
     6661 cgatttagtg ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt
     6721 agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt
     6781 aatagtggac tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt
     6841 gatttataag ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa
     6901 aaatttaacg cgaattttaa caaaatatta acgcttacaa tttag
//
