LOCUS       Exported                6526 bp ds-DNA     circular SYN 06-MAR-2024
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6526)
  AUTHORS   Institute for Molecular Bioscience
  TITLE     Direct Submission
  JOURNAL   Exported Friday, Nov 8, 2024 from SnapGene 5.0.8
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6526
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     misc_feature    769..789
                     /label=SV_CMV Forward
     misc_feature    complement(798..818)
                     /label=SV CMV Reverse
     misc_feature    863..882
                     /label=SV_T7
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             966..1682
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             966..1682
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     regulatory      966..969
                     /regulatory_class="other"
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     primer_bind     complement(1011..1032)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1272..1291)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1619..1640
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    1683..2090
                     /label=3XFP4 cyto
     primer_bind     complement(2117..2134)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2123..2347
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     misc_feature    2123..2134
                     /label=BGH FW
     misc_feature    complement(2329..2346)
                     /label=SV BGH REV
     rep_origin      2393..2821
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     misc_feature    2399..2417
                     /label=SV FL ORI Forward
     misc_feature    complement(2693..2713)
                     /label=SV FL ORI Reverse
     promoter        2835..3164
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     misc_feature    2843..2863
                     /label=SV SV40 PROMOTER Forward
     rep_origin      3015..3150
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3231..4025
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     misc_feature    3616..3633
                     /label=SV NeoR/KanR Forward
     misc_feature    complement(3693..3712)
                     /label=SV NeoR/KanR Reverse
     polyA_signal    4199..4320
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     misc_feature    4199..4220
                     /label=SV SV40pA-Forward
     misc_feature    complement(4236..4255)
                     /label=SV_SV40pA-R
     primer_bind     complement(4369..4385)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_feature    complement(4382..4404)
                     /label=M13_pUC RV
     protein_bind    4393..4409
                     /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)."
     promoter        complement(4417..4447)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4462..4483
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     misc_feature    4462..4483
                     /label=SV CAP binding Forward
     rep_origin      complement(4771..5359)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     misc_feature    complement(4851..4870)
                     /label=pBR322ori-F
     misc_feature    4851..4870
                     /label=SV ORI FORWARD
     misc_feature    5052..5070
                     /label=SV ORI MID FORWARD
     CDS             complement(5530..6390)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     misc_feature    complement(5534..5554)
                     /label=SV Amp 5' Reverse
     misc_feature    6084..6103
                     /label=Amp_mid FW
     misc_feature    complement(6215..6235)
                     
     promoter        complement(6391..6495)
                     /gene="bla"
                     /label=AmpR promoter
     misc_feature    6396..6418
                     /label=SV AmpR Promoter Forward 2
     misc_feature    complement(6472..6492)
                     /label=SV AmpR Promoter Reverse 2
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc
      901 gtttaaacgg gccctctaga ctcgagcggc cgccactgtg ctggatatct gcagaattcg
      961 ccaccatggt gagcaagggc gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc
     1021 tggacggcga cgtaaacggc cacaagttca gcgtgtccgg cgagggcgag ggcgatgcca
     1081 cctacggcaa gctgaccctg aagttcatct gcaccaccgg caagctgccc gtgccctggc
     1141 ccaccctcgt gaccaccctg acctacggcg tgcagtgctt cagccgctac cccgaccaca
     1201 tgaagcagca cgacttcttc aagtccgcca tgcccgaagg ctacgtccag gagcgcacca
     1261 tcttcttcaa ggacgacggc aactacaaga cccgcgccga ggtgaagttc gagggcgaca
     1321 ccctggtgaa ccgcatcgag ctgaagggca tcgacttcaa ggaggacggc aacatcctgg
     1381 ggcacaagct ggagtacaac tacaacagcc acaacgtcta tatcatggcc gacaagcaga
     1441 agaacggcat caaggtgaac ttcaagatcc gccacaacat cgaggacggc agcgtgcagc
     1501 tcgccgacca ctaccagcag aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca
     1561 accactacct gagcacccag tccgccctga gcaaagaccc caacgagaag cgcgatcaca
     1621 tggtcctgct ggagttcgtg accgccgccg ggatcactct cggcatggac gagctgtaca
     1681 agtccggact cagatctatc gtagcttcgg acttcccgcc accacctacg gatgaagagt
     1741 taagacttgc tttgccagag acaccaatgc ttcttggttt taatgctcct gctacatcag
     1801 aaccgagctc attcgaattt ccaccaccac ctacggatga agagttaaga cttgctttgc
     1861 cagagacgcc aatgcttctt ggttttaatg ctcctgctac atcggaaccg agctcgttcg
     1921 aatttccacc gcctccaaca gaagatgaac tagaaatcat ccgggaaaca gcatcctcgc
     1981 tagattctag ttttacaaga ggggatttag ctagtttgag aaatgctatt aatcgccata
     2041 gtcaaaattt ctctgatttc ccaccaatcc caacagaaga agagttgtaa aagcttaagt
     2101 ttaaaccgct gatcagcctc gactgtgcct tctagttgcc agccatctgt tgtttgcccc
     2161 tcccccgtgc cttccttgac cctggaaggt gccactccca ctgtcctttc ctaataaaat
     2221 gaggaaattg catcgcattg tctgagtagg tgtcattcta ttctgggggg tggggtgggg
     2281 caggacagca agggggagga ttgggaagac aatagcaggc atgctgggga tgcggtgggc
     2341 tctatggctt ctgaggcgga aagaaccagc tggggctcta gggggtatcc ccacgcgccc
     2401 tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt
     2461 gccagcgccc tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc
     2521 ggctttcccc gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta
     2581 cggcacctcg accccaaaaa acttgattag ggtgatggtt cacgtagtgg gccatcgccc
     2641 tgatagacgg tttttcgccc tttgacgttg gagtccacgt tctttaatag tggactcttg
     2701 ttccaaactg gaacaacact caaccctatc tcggtctatt cttttgattt ataagggatt
     2761 ttgccgattt cggcctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat
     2821 taattctgtg gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc ccagcaggca
     2881 gaagtatgca aagcatgcat ctcaattagt cagcaaccag gtgtggaaag tccccaggct
     2941 ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc atagtcccgc
     3001 ccctaactcc gcccatcccg cccctaactc cgcccagttc cgcccattct ccgccccatg
     3061 gctgactaat tttttttatt tatgcagagg ccgaggccgc ctctgcctct gagctattcc
     3121 agaagtagtg aggaggcttt tttggaggcc taggcttttg caaaaagctc ccgggagctt
     3181 gtatatccat tttcggatct gatcaagaga caggatgagg atcgtttcgc atgattgaac
     3241 aagatggatt gcacgcaggt tctccggccg cttgggtgga gaggctattc ggctatgact
     3301 gggcacaaca gacaatcggc tgctctgatg ccgccgtgtt ccggctgtca gcgcaggggc
     3361 gcccggttct ttttgtcaag accgacctgt ccggtgccct gaatgaactg caggacgagg
     3421 cagcgcggct atcgtggctg gccacgacgg gcgttccttg cgcagctgtg ctcgacgttg
     3481 tcactgaagc gggaagggac tggctgctat tgggcgaagt gccggggcag gatctcctgt
     3541 catctcacct tgctcctgcc gagaaagtat ccatcatggc tgatgcaatg cggcggctgc
     3601 atacgcttga tccggctacc tgcccattcg accaccaagc gaaacatcgc atcgagcgag
     3661 cacgtactcg gatggaagcc ggtcttgtcg atcaggatga tctggacgaa gagcatcagg
     3721 ggctcgcgcc agccgaactg ttcgccaggc tcaaggcgcg catgcccgac ggcgaggatc
     3781 tcgtcgtgac ccatggcgat gcctgcttgc cgaatatcat ggtggaaaat ggccgctttt
     3841 ctggattcat cgactgtggc cggctgggtg tggcggaccg ctatcaggac atagcgttgg
     3901 ctacccgtga tattgctgaa gagcttggcg gcgaatgggc tgaccgcttc ctcgtgcttt
     3961 acggtatcgc cgctcccgat tcgcagcgca tcgccttcta tcgccttctt gacgagttct
     4021 tctgagcggg actctggggt tcgaaatgac cgaccaagcg acgcccaacc tgccatcacg
     4081 agatttcgat tccaccgccg ccttctatga aaggttgggc ttcggaatcg ttttccggga
     4141 cgccggctgg atgatcctcc agcgcgggga tctcatgctg gagttcttcg cccaccccaa
     4201 cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa atttcacaaa
     4261 taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca atgtatctta
     4321 tcatgtctgt ataccgtcga cctctagcta gagcttggcg taatcatggt catagctgtt
     4381 tcctgtgtga aattgttatc cgctcacaat tccacacaac atacgagccg gaagcataaa
     4441 gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca ttaattgcgt tgcgctcact
     4501 gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc
     4561 ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg
     4621 ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc
     4681 cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag
     4741 gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca
     4801 tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca
     4861 ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg
     4921 atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag
     4981 gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt
     5041 tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca
     5101 cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg
     5161 cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa gaacagtatt
     5221 tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc
     5281 cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg
     5341 cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg
     5401 gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta
     5461 gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg
     5521 gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg
     5581 ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc
     5641 atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc cagatttatc
     5701 agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc
     5761 ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag
     5821 tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat
     5881 ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg
     5941 caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt
     6001 gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag
     6061 atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg
     6121 accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt
     6181 aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct
     6241 gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac
     6301 tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat
     6361 aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat
     6421 ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca
     6481 aataggggtt ccgcgcacat ttccccgaaa agtgccacct gacgtc
//
