LOCUS       Exported                6720 bp ds-DNA     circular SYN 25-JUN-2019
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6720)
  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..6720
                     /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     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..2744
                     /codon_start=1
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             2745..2786
                     /codon_start=1
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             2787..2828
                     /codon_start=1
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     primer_bind     2827..2854
                     /label=TOL784_Traptavidin_F
     CDS             2832..3215
                     /codon_start=1
                     /label=Traptavidin?
                     /translation="MAEAGITGTWYNQLGSTFIVTAGADGALTGTYESAVGNAEGDYVL
                     TGRYDSAPATDGSGTALGWTVAWKNNYRNAHSATTWSGQYVGGAEARINTQWLLTSGTT
                     EANAWKSTLVGHDTFTKVKPSAAS"
     primer_bind     complement(3191..3235)
                     /label=TOL787_Traptavidin_R2
     primer_bind     3196..3233
                     /label=TOL786_linker_p65HSF1_F3
     primer_bind     3215..3253
                     /label=TOL35
     CDS             3216..3269
                     /codon_start=1
                     /label=Linker
                     /translation="SAGGGGSGGGGSGGGGSG"
     primer_bind     3216..3235
                     /label=TOL774_linker_p65HSF1_F
     primer_bind     3238..3256
                     /label=TOL418_GB1linker_seq_F
     primer_bind     complement(3253..3269)
                     /label=TOL316_GCN4_p65HSF1_R2
     primer_bind     complement(3257..3278)
                     /label=TOL397
     primer_bind     complement(3257..3278)
                     /label=TOL-29
     primer_bind     complement(3260..3278)
                     /label=TOL405
     CDS             3279..3911
                     /codon_start=1
                     /label=DNMT3A
                     /translation="PSRLQMFFANNHDQEFDPPKVYPPVPAEKRKPIRVLSLFDGIATG
                     LLVLKDLGIQVDRYIASEVCEDSITVGMVRHQGKIMYVGDVRSVTQKHIQEWGPFDLVI
                     GGSPCNDLSIVNPARKGLYEGTGRLFFEFYRLLHDARPKEGDDRPFFWLFENVVAMGVS
                     DKRDISRFLESNPVMIDAKEVSAAHRARYFWGNLPGMNRPLASTVNDK"
     misc_feature    3279..3911
                     /label=Amino acid translation
     primer_bind     complement(3885..3911)
                     /label=TOL403_DNMT3a_codon_R
     misc_feature    3912..4211
                     /label=DNMT3a Codon Change
     primer_bind     3930..3949
                     /label=TOL453_DNMT3a_qPCR1_F
     primer_bind     complement(4008..4027)
                     /label=TOL454_DNMT3a_qPCR1_R
     misc_feature    complement(4215..4238)
                     /label=UPS3
     polyA_signal    4239..4477
                     /label=SV40 polyA
     primer_bind     4239..4278
                     /label=TOL400_UPS_polyA_F
     primer_bind     complement(4299..4323)
                     /label=TOL419_SV40_seq_R
     rep_origin      4956..5575
                     /label=pBR322 Orgin
     CDS             complement(5730..6590)
                     /codon_start=1
                     /label=Ampicillin
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(6632..6660)
                     /label=AMP promoter
     primer_bind     join(6703..6720,1..25)
                     /label=TOL838_GFP_bGHpolyA_R
     primer_bind     complement(join(6705..6720,1..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 tgggtaagcc tatccctaac cctctcctcg gtctcgattc tacgggcaag cccatcccca
     2761 accccctgct gggcctggac agcaccggta aacctattcc taatccactc ctcggactcg
     2821 attccacaca tatggctgaa gctggtatca ccggcacctg gtacaaccag ctgggatcca
     2881 ccttcatcgt taccgctggt gctgacggtg ctctgaccgg tacctacgaa tccgctgttg
     2941 gtaacgctga aggcgattac gttctgaccg gtcgttacga ctccgctccg gctaccgacg
     3001 gttccggaac cgctctgggt tggaccgttg cttggaaaaa caactaccgt aacgctcact
     3061 ccgctaccac ctggtctggc cagtacgttg gtggtgctga agctcgtatc aacacccagt
     3121 ggttgttgac ctccggcacc accgaagcta acgcgtggaa atccaccctg gttggtcacg
     3181 acaccttcac caaagttaaa ccgtccgctg cttccagcgc tggagggggt ggctccggcg
     3241 gaggaggctc tggtggcgga gggagtggag cggccgctcc ctcccggctc cagatgttct
     3301 tcgctaataa ccacgaccag gaatttgacc ctccaaaggt ttacccacct gtcccagctg
     3361 agaagaggaa gcccatccgg gtgctgtctc tctttgatgg aatcgctaca gggctcctgg
     3421 tgctgaagga cttgggcatt caggtggacc gctacattgc ctcggaggtg tgtgaggact
     3481 ccatcacggt gggcatggtg cggcaccagg ggaagatcat gtacgtcggg gacgtccgca
     3541 gcgtcacaca gaagcatatc caggagtggg gcccattcga tctggtgatt gggggcagtc
     3601 cctgcaatga cctctccatc gtcaaccctg ctcgcaaggg cctctacgag ggcactggcc
     3661 ggctgttctt tgagttctac cgcctcctgc atgatgcgcg gcccaaggag ggagatgatc
     3721 gccccttctt ctggctcttt gagaatgtgg tggccatggg cgttagtgac aagagggaca
     3781 tctcgcgatt tctcgagtcc aaccctgtga tgattgatgc caaagaagtg tcagctgcac
     3841 acagggcccg ctacttctgg ggtaaccttc ccggtatgaa caggccgttg gcatccactg
     3901 tgaatgataa gcttgagttg caagagtgcc ttgaacacgg acggatagcc aaattctcaa
     3961 aggtacgaac tataacaact cggtccaact caataaagca aggtaaggac cagcattttc
     4021 ccgtgtttat gaacgaaaaa gaagacattt tgtggtgtac cgagatggaa agggtcttcg
     4081 gtttcccagt gcattatact gatgttagca acatgagtag gttggcccgg caacggctgt
     4141 tgggtaggag ttggtcagtc cccgttatcc gccacctttt cgcgccgttg aaagagtact
     4201 tcgcctgcgt atgacgttga gtctctgttt gtggtgactg atcataatca gccataccac
     4261 atttgtagag gttttacttg ctttaaaaaa cctcccacac ctccccctga acctgaaaca
     4321 taaaatgaat gcaattgttg ttgttaactt gtttattgca gcttataatg gttacaaata
     4381 aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg
     4441 tttgtccaaa ctcatcaatg tatcttaatg cgtcgagtgc attctagttg tggtttgtcc
     4501 aaactcatca atgtatctta tcatgtctgt ataccgtcga cctctagcta gagcttggcg
     4561 taatcatggt catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac
     4621 atacgagccg gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca
     4681 ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat
     4741 taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc
     4801 tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca
     4861 aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca
     4921 aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg
     4981 ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg
     5041 acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt
     5101 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt
     5161 tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc
     5221 tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt
     5281 gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt
     5341 agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc
     5401 tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa
     5461 agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt
     5521 tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct
     5581 acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta
     5641 tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa
     5701 agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc
     5761 tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact
     5821 acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc
     5881 tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt
     5941 ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta
     6001 agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg
     6061 tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt
     6121 acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc
     6181 agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt
     6241 actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc
     6301 tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc
     6361 gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa
     6421 ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac
     6481 tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa
     6541 aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt
     6601 tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa
     6661 tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct
//
