LOCUS       Exported                6555 bp ds-DNA     circular SYN 18-JUN-2019
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6555)
  AUTHORS   Ryan Lister
  TITLE     Direct Submission
  JOURNAL   Exported Tuesday, Oct 31, 2023 from SnapGene 5.0.8
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6555
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     polyA_signal    6..230
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     6..27
                     /label=OC-310_pLenti_1_dCas9_5.F
     primer_bind     complement(204..230)
                     /label=OC-311_pLenti_1_dCas9_5.R
     primer_bind     complement(210..253)
                     /label=TOL827_pGHpolyA_R
     primer_bind     211..254
                     /label=TOL825_pEF1a-GFP_R
     CDS             complement(240..956)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     240..260
                     /label=TOL195_GFP_PUF_polyA_R
     primer_bind     complement(666..684)
                     /label=TOL102_GFP_backbone_F
     primer_bind     682..696
                     /label=TOL103_GFP_backbone_R
     primer_bind     complement(939..956)
                     /label=TOL194_T2A_GFP_F
     primer_bind     complement(949..988)
                     /label=TOL837_GFP_bGHpolyA_F
     primer_bind     949..987
                     /label=TOL840_pSV40_R
     promoter        complement(981..1191)
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      987..1122
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(1170..1224)
                     /label=TOL839_pSV40_F
     primer_bind     1183..1225
                     /label=TOL847_pEF1a_F
     primer_bind     complement(1212..1229)
                     /label=TOL847_pEF1a_F
     primer_bind     1213..1229
                     /label=TOL839_pSV40_F
     primer_bind     1291..1312
                     /label=TOL318_GCN4_pEF1a_F
     primer_bind     complement(1291..1312)
                     /label=TOL319_GCN4_pEF1a_R
     primer_bind     1331..1356
                     /label=OL-D206
     promoter        1406..2663
                     /label=EF1a
     primer_bind     complement(1478..1501)
                     /label=OL-D243
     primer_bind     1846..1867
                     /label=OL -D154
     primer_bind     2458..2479
                     /label=OL- D155
     primer_bind     2524..2545
                     /label=TOL417_GCN4_seq_F
     primer_bind     complement(2655..2674)
                     /label=TOL848_pEF1a_R
     misc_feature    2670..2675
                     
     primer_bind     complement(2673..2702)
                     /label=TOL401_3xTy1_GCN4_ R
     CDS             2676..2678
                     /codon_start=1
                     /label=ATG
                     /translation="M"
     CDS             2682..2702
                     /codon_start=1
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2703..2774
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags"
                     /label=3xFLAG
                     /translation="DYKDDDDKDYKDDDDKDYKDDDDK"
     primer_bind     2774..2814
                     /label=TOL773_SpyCatcher_R
     CDS             2775..3050
                     /codon_start=1
                     /label=SpyCatcher
                     /translation="DSATHIKFSKRDEDGKELAGATMELRDSSGKTISTWISDGQVKDF
                     YLYPGKYTFVETAAPDGYEVATAITFTVNEQGQVTVNGKATKGDAHI"
     primer_bind     complement(3013..3070)
                     /label=TOL775_SpyCatcher_F2
     primer_bind     3031..3070
                     /label=TOL774_linker_p65HSF1_F
     CDS             3051..3104
                     /codon_start=1
                     /label=Linker
                     /translation="SAGGGGSGGGGSGGGGSG"
     primer_bind     3051..3088
                     /label=TOL35
     primer_bind     3051..3068
                     /label=TOL786_linker_p65HSF1_F3
     primer_bind     3051..3067
                     /label=TOL780_linker_p65HSF1_F2
     primer_bind     3073..3091
                     /label=TOL418_GB1linker_seq_F
     primer_bind     complement(3088..3104)
                     /label=TOL316_GCN4_p65HSF1_R2
     primer_bind     complement(3092..3113)
                     /label=TOL397
     primer_bind     complement(3092..3113)
                     /label=TOL-29
     primer_bind     complement(3095..3113)
                     /label=TOL405
     CDS             3114..3746
                     /codon_start=1
                     /label=DNMT3A
                     /translation="PSRLQMFFANNHDQEFDPPKVYPPVPAEKRKPIRVLSLFDGIATG
                     LLVLKDLGIQVDRYIASEVCEDSITVGMVRHQGKIMYVGDVRSVTQKHIQEWGPFDLVI
                     GGSPCNDLSIVNPARKGLYEGTGRLFFEFYRLLHDARPKEGDDRPFFWLFENVVAMGVS
                     DKRDISRFLESNPVMIDAKEVSAAHRARYFWGNLPGMNRPLASTVNDK"
     misc_feature    3114..3746
                     /label=Amino acid translation
     primer_bind     complement(3720..3746)
                     /label=TOL403_DNMT3a_codon_R
     misc_feature    3747..4046
                     /label=DNMT3a Codon Change
     primer_bind     3765..3784
                     /label=TOL453_DNMT3a_qPCR1_F
     primer_bind     complement(3843..3862)
                     /label=TOL454_DNMT3a_qPCR1_R
     misc_feature    complement(4050..4073)
                     /label=UPS3
     polyA_signal    4074..4312
                     /label=SV40 polyA
     primer_bind     4074..4113
                     /label=TOL400_UPS_polyA_F
     primer_bind     complement(4134..4158)
                     /label=TOL419_SV40_seq_R
     rep_origin      4791..5410
                     /label=pBR322 Orgin
     CDS             complement(5565..6425)
                     /codon_start=1
                     /label=Ampicillin
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(6467..6495)
                     /label=AMP promoter
     primer_bind     join(6538..6555,1..25)
                     /label=TOL838_GFP_bGHpolyA_R
     primer_bind     complement(join(6540..6555,1..25))
                     /label=TOL846_p65HSF1_R
ORIGIN
        1 gacgtccata gagcccaccg catccccagc atgcctgcta ttgtcttccc aatcctcccc
       61 cttgctgtcc tgccccaccc caccccccag aatagaatga cacctactca gacaatgcga
      121 tgcaatttcc tcattttatt aggaaaggac agtgggagtg gcaccttcca gggtcaagga
      181 aggcacgggg gaggggcaaa caacagatgg ctggcaacta gaaggcacag tcaactgccc
      241 ttgtacagct cgtccatgcc gagagtgatc ccggcggcgg tcacgaactc cagcaggacc
      301 atgtgatcgc gcttctcgtt ggggtctttg ctcagggcgg actgggtgct caggtagtgg
      361 ttgtcgggca gcagcacggg gccgtcgccg atgggggtgt tctgctggta gtggtcggcg
      421 agctgcacgc tgccgtcctc gatgttgtgg cggatcttga agttcacctt gatgccgttc
      481 ttctgcttgt cggccatgat atagacgttg tggctgttgt agttgtactc cagcttgtgc
      541 cccaggatgt tgccgtcctc cttgaagtcg atgcccttca gctcgatgcg gttcaccagg
      601 gtgtcgccct cgaacttcac ctcggcgcgg gtcttgtagt tgccgtcgtc cttgaagaag
      661 atggtgcgct cctggacgta gccttcgggc atggcggact tgaagaagtc gtgctgcttc
      721 atgtggtcgg ggtagcggct gaagcactgc acgccgtagg tcagggtggt cacgagggtg
      781 ggccagggca cgggcagctt gccggtggtg cagatgaact tcagggtcag cttgccgtag
      841 gtggcatcgc cctcgccctc gccggacacg ctgaacttgt ggccgtttac gtcgccgtcc
      901 agctcgacca ggatgggcac caccccggtg aacagctcct cgcccttgct caccatggtg
      961 gccgtacggg tggcgcgcgt agcctaggcc tccaaaaaag cctcctcact acttctggaa
     1021 tagctcagag gcagaggcgg cctcggcctc tgcataaata aaaaaaatta gtcagccatg
     1081 gggcggagaa tgggcggaac tgggcggagt taggggcggg atgggcggag ttaggggcgg
     1141 gactatggtt gctgactaat tgagatgcat gctttgcata cttctgcctg cctgaattca
     1201 cgtcgacgga tcgggagatc tcccgatccc ctatggtgca ctctcagtac aatctgctct
     1261 gatgccgcat agttaagcca gtatctgctc cctgcttgtg tgttggaggt cgctgagtag
     1321 tgcgcgagca aaatttaagc tacaacaagg caaggcttga ccgacaattg catgaagaat
     1381 ctgcttaggg ttaggcgttt tgcgtgcaaa gatggataaa gttttaaaca gagaggaatc
     1441 tttgcagcta atggaccttc taggtcttga aaggagtggg aattggctcc ggtgcccgtc
     1501 agtgggcaga gcgcacatcg cccacagtcc ccgagaagtt ggggggaggg gtcggcaatt
     1561 gaaccggtgc ctagagaagg tggcgcgggg taaactggga aagtgatgtc gtgtactggc
     1621 tccgcctttt tcccgagggt gggggagaac cgtatataag tgcagtagtc gccgtgaacg
     1681 ttctttttcg caacgggttt gccgccagaa cacaggtaag tgccgtgtgt ggttcccgcg
     1741 ggcctggcct ctttacgggt tatggccctt gcgtgccttg aattacttcc acctggctgc
     1801 agtacgtgat tcttgatccc gagcttcggg ttggaagtgg gtgggagagt tcgaggcctt
     1861 gcgcttaagg agccccttcg cctcgtgctt gagttgaggc ctggcctggg cgctggggcc
     1921 gccgcgtgcg aatctggtgg caccttcgcg cctgtctcgc tgctttcgat aagtctctag
     1981 ccatttaaaa tttttgatga cctgctgcga cgcttttttt ctggcaagat agtcttgtaa
     2041 atgcgggcca agatctgcac actggtattt cggtttttgg ggccgcgggc ggcgacgggg
     2101 cccgtgcgtc ccagcgcaca tgttcggcga ggcggggcct gcgagcgcgg ccaccgagaa
     2161 tcggacgggg gtagtctcaa gctggccggc ctgctctggt gcctggcctc gcgccgccgt
     2221 gtatcgcccc gccctgggcg gcaaggctgg cccggtcggc accagttgcg tgagcggaaa
     2281 gatggccgct tcccggccct gctgcaggga gctcaaaatg gaggacgcgg cgctcgggag
     2341 agcgggcggg tgagtcaccc acacaaagga aaagggcctt tccgtcctca gccgtcgctt
     2401 catgtgactc cacggagtac cgggcgccgt ccaggcacct cgattagttc tcgagctttt
     2461 ggagtacgtc gtctttaggt tggggggagg ggttttatgc gatggagttt ccccacactg
     2521 agtgggtgga gactgaagtt aggccagctt ggcacttgat gtaattctcc ttggaatttg
     2581 ccctttttga gtttggatct tggttcattc tcaagcctca gacagtggtt caaagttttt
     2641 ttcttccatt tcaggtgtcg tgacgtacgg ccaccatggg tcctaagaaa aagaggaagg
     2701 tggattacaa ggatgacgac gataaggact ataaggacga tgatgacaag gactacaaag
     2761 atgatgacga taaagatagt gctacccata ttaaattctc aaaacgtgat gaggacggca
     2821 aagagttagc tggtgcaact atggagttgc gtgattcatc tggtaaaact attagtacat
     2881 ggatttcaga tggacaagtg aaagatttct acctgtatcc aggaaaatat acatttgtcg
     2941 aaaccgcagc accagacggt tatgaggtag caactgctat tacctttaca gttaatgagc
     3001 aaggtcaggt tactgtaaat ggcaaagcaa ctaaaggtga cgctcatatt agcgctggag
     3061 ggggtggctc cggcggagga ggctctggtg gcggagggag tggagcggcc gctccctccc
     3121 ggctccagat gttcttcgct aataaccacg accaggaatt tgaccctcca aaggtttacc
     3181 cacctgtccc agctgagaag aggaagccca tccgggtgct gtctctcttt gatggaatcg
     3241 ctacagggct cctggtgctg aaggacttgg gcattcaggt ggaccgctac attgcctcgg
     3301 aggtgtgtga ggactccatc acggtgggca tggtgcggca ccaggggaag atcatgtacg
     3361 tcggggacgt ccgcagcgtc acacagaagc atatccagga gtggggccca ttcgatctgg
     3421 tgattggggg cagtccctgc aatgacctct ccatcgtcaa ccctgctcgc aagggcctct
     3481 acgagggcac tggccggctg ttctttgagt tctaccgcct cctgcatgat gcgcggccca
     3541 aggagggaga tgatcgcccc ttcttctggc tctttgagaa tgtggtggcc atgggcgtta
     3601 gtgacaagag ggacatctcg cgatttctcg agtccaaccc tgtgatgatt gatgccaaag
     3661 aagtgtcagc tgcacacagg gcccgctact tctggggtaa ccttcccggt atgaacaggc
     3721 cgttggcatc cactgtgaat gataagcttg agttgcaaga gtgccttgaa cacggacgga
     3781 tagccaaatt ctcaaaggta cgaactataa caactcggtc caactcaata aagcaaggta
     3841 aggaccagca ttttcccgtg tttatgaacg aaaaagaaga cattttgtgg tgtaccgaga
     3901 tggaaagggt cttcggtttc ccagtgcatt atactgatgt tagcaacatg agtaggttgg
     3961 cccggcaacg gctgttgggt aggagttggt cagtccccgt tatccgccac cttttcgcgc
     4021 cgttgaaaga gtacttcgcc tgcgtatgac gttgagtctc tgtttgtggt gactgatcat
     4081 aatcagccat accacatttg tagaggtttt acttgcttta aaaaacctcc cacacctccc
     4141 cctgaacctg aaacataaaa tgaatgcaat tgttgttgtt aacttgttta ttgcagctta
     4201 taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat ttttttcact
     4261 gcattctagt tgtggtttgt ccaaactcat caatgtatct taatgcgtcg agtgcattct
     4321 agttgtggtt tgtccaaact catcaatgta tcttatcatg tctgtatacc gtcgacctct
     4381 agctagagct tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc
     4441 acaattccac acaacatacg agccggaagc ataaagtgta aagcctgggg tgcctaatga
     4501 gtgagctaac tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg
     4561 tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg
     4621 cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg
     4681 gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga
     4741 aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg
     4801 gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag
     4861 aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc
     4921 gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg
     4981 ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt
     5041 cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc
     5101 ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc
     5161 actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg
     5221 tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca
     5281 gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc
     5341 ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat
     5401 cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt
     5461 ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt
     5521 tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc
     5581 agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc
     5641 gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata
     5701 ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg
     5761 gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc
     5821 cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct
     5881 acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa
     5941 cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt
     6001 cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca
     6061 ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac
     6121 tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca
     6181 atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt
     6241 tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc
     6301 actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca
     6361 aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata
     6421 ctcatactct tcctttttca atattattga agcatttatc agggttattg tctcatgagc
     6481 ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc
     6541 cgaaaagtgc cacct
//
