LOCUS       sequence-386596-        6150 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  ""pEF1s(orig)"" reporter plasmid.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6150)
  AUTHORS   Ma Y, Budde MW, Zhu J, Elowitz MB
  TITLE     Tuning Methylation-Dependent Silencing Dynamics by Synthetic 
            Modulation of CpG Density.
  JOURNAL   ACS Synth Biol. 2023 Aug 12. doi: 10.1021/acssynbio.3c00078.
  PUBMED    37572041
REFERENCE   2  (bases 1 to 6150)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "ACS Synth 
            Biol. 2023 Aug 12. doi: 10.1021/acssynbio.3c00078."
COMMENT     Sequence Label: vYM004_pNeo_phiC31_nTRE_pEF1s(orig)_citrine
FEATURES             Location/Qualifiers
     source          1..6150
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      77..665
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     566..585
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     polyA_signal    849..983
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     polyA_signal    1086..1207
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     protein_bind    1244..1313
                     /label=attB
                     /bound_moiety="phage phi-C31 integrase"
                     /note="attB site for the phi-C31 integrase (Groth et al., 
                     2000)"
     CDS             1579..2382
                     /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="MGSAIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQ
                     GRPVLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQD
                     LLSSHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDE
                     EHQGLAPAELFARLKARMPDGDDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQ
                     DIALATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(1642..1661)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2252..2271
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(2414..2431)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2420..2644
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     protein_bind    2717..2735
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2753..2771
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2789..2807
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2825..2843
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2861..2879
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     promoter        2947..3158
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     LTR             3171..3439
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from human 
                     T-cell leukemia virus (HTLV) type 1"
     CDS             3523..3900
                     /codon_start=1
                     /gene="HIST1H2BJ"
                     /product="human histone H2B"
                     /label=H2B
                     /translation="MPEPAKSAPAPKKGSKKAVTKAQKKGGKKRKRSRKESYSIYVYKV
                     LKQVHPDTGISSKAMGIMNSFVNDIFERIAGEASRLAHYNKRSTITSREIQTAVRLLLP
                     GELAKHAVSEGTKAITKYTSAK"
     regulatory      3919..3928
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(3925..3927,3928..3930,3931..4641)
                     /codon_start=1
                     /product="enhanced variant of YFP (Heikal et al., 2001)"
                     /label=Citrine
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLMCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(3970..3991)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(4231..4250)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4578..4599
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(4692..4711)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4693..4711)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     polyA_signal    4985..5106
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     promoter        5132..5195
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5196..5264,5265..6056)
                     /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"
     primer_bind     complement(5414..5433)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa
       61 agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa
      121 aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc
      181 cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt
      241 agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc
      301 tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac
      361 gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca
      421 gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg
      481 ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag
      541 gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt
      601 ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat
      661 ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc
      721 acatgttctt tcctgcggta ccgatcataa tcagccatac cacatttgta gaggttttac
      781 ttgctttaaa aaacctccca cacctccccc tgaacctgaa acataaaatg aatgcaattg
      841 ttgttgttaa cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa
      901 atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca
      961 atgtatctta tcatgtctgg atcataatca gccataccac atttgtagag gttttacttg
     1021 ctttaaaaaa cctcccacac ctccccctga acctgaaaca taaaatgaat gcaattgttg
     1081 ttgttaactt gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt
     1141 tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg
     1201 tatcttatca tgtctgctcg aggtcgacga tgtaggtcac ggtctcgaag ccgcggtgcg
     1261 ggtgccaggg cgtgcccttg ggctccccgg gcgcgtactc cacctcaccc atctggtcca
     1321 tcatgatgaa cgggtcgagg tggcggtagt tgatcccggc gaacgcgcgg cgcaccggga
     1381 agccctcgcc ctcgaaaccg ctgggcgcgg tggtcacggt gagcacggga cgtgcgacgg
     1441 cgtcggcggg tgcggatacg cggggcagcg tcagcgggtt ctcgacggtc acggcgggca
     1501 tgtcgacggt atcgataagc ttacgcgtgc tagcgcatgc agatctacta gtcggccgga
     1561 taaagatatc aactcgagat gggatcggcc attgaacaag atggattgca cgcaggttct
     1621 ccggccgctt gggtggagag gctattcggc tatgactggg cacaacagac aatcggctgc
     1681 tctgatgccg ccgtgttccg gctgtcagcg caggggcgcc cggttctttt tgtcaagacc
     1741 gacctgtccg gtgccctgaa tgaactgcag gacgaggcag cgcggctatc gtggctggcc
     1801 acgacgggcg ttccttgcgc agctgtgctc gacgttgtca ctgaagcggg aagggactgg
     1861 ctgctattgg gcgaagtgcc ggggcaggat ctcctgtcat ctcaccttgc tcctgccgag
     1921 aaagtatcca tcatggctga tgcaatgcgg cggctgcata cgcttgatcc ggctacctgc
     1981 ccattcgacc accaagcgaa acatcgcatc gagcgagcac gtactcggat ggaagccggt
     2041 cttgtcgatc aggatgatct ggacgaagag catcaggggc tcgcgccagc cgaactgttc
     2101 gccaggctca aggcgcgcat gcccgacggc gatgatctcg tcgtgaccca tggcgatgcc
     2161 tgcttgccga atatcatggt ggaaaatggc cgcttttctg gattcatcga ctgtggccgg
     2221 ctgggtgtgg cggaccgcta tcaggacata gcgttggcta cccgtgatat tgctgaagag
     2281 cttggcggcg aatgggctga ccgcttcctc gtgctttacg gtatcgccgc tcccgattcg
     2341 cagcgcatcg ccttctatcg ccttcttgac gagttcttct gaggggatca attctctaga
     2401 gctcgctgat cagcctcgac tgtgccttct agttgccagc catctgttgt ttgcccctcc
     2461 cccgtgcctt ccttgaccct ggaaggtgcc actcccactg tcctttccta ataaaatgag
     2521 gaaattgcat cgcattgtct gagtaggtgt cattctattc tggggggtgg ggtggggcag
     2581 gacagcaagg gggaggattg ggaagagaat agcaggcatg ctggggatgc ggtgggctct
     2641 atggcttctg aggcggaaag aaccagctgg gggctagcaa agcctaggaa cagctgcata
     2701 tagcactgca gtttactccc tatcagtgat agagagtcaa aaggccatgc cctccctatc
     2761 agtgatagag agagcagact caaggtggtc cctatcagtg atagagagat tgagcaagag
     2821 ggcctcccta tcagtgatag agattgatcc agtcagggcc tccctatcag tgatagagaa
     2881 gctatgtgca ggtagggctg tacagtggga atgcatggat ctgcgatcgc tccggtgccc
     2941 gtcagtgggc agagcgcaca tcgcccacag tccccgagaa gttgggggga ggggtcggca
     3001 attgaaccgg tgcctagaga aggtggcgcg gggtaaactg ggaaagtgat gtcgtgtact
     3061 ggctccgcct ttttcccgag ggtgggggag aaccgtatat aagtgcagta gtcgccgtga
     3121 acgttctttt tcgcaacggg tttgccgcca gaacacagct gaagcttcga ggggctcgca
     3181 tctctccttc acgcgcccgc cgccctacct gaggccgcca tccacgccgg ttgagtcgcg
     3241 ttctgccgcc tcccgcctgt ggtgcctcct gaactgcgtc cgccgtctag gtaagtttaa
     3301 agctcaggtc gagaccgggc ctttgtccgg cgctcccttg gagcctacct agactcagcc
     3361 ggctctccac gctttgcctg accctgcttg ctcaactcta cgtctttgtt tcgttttctg
     3421 ttctgcgccg ttacagatcc aagctgtgac cggcgcctac gcggccgctc tagcccgggc
     3481 ggatcccccg ggctgcagga attcgatatc aagcttgcca ccatgccaga gccagcgaag
     3541 tctgctcccg ccccgaaaaa gggctccaag aaggcggtga ctaaggcgca gaagaaaggc
     3601 ggcaagaagc gcaagcgcag ccgcaaggag agctattcca tctatgtgta caaggttctg
     3661 aagcaggtcc accctgacac cggcatttcg tccaaggcca tgggcatcat gaattcgttt
     3721 gtgaacgaca ttttcgagcg catcgcaggt gaggcttccc gcctggcgca ttacaacaag
     3781 cgctcgacca tcacctccag ggagatccag acggccgtgc gcctgctgct gcctggggag
     3841 ttggccaagc acgccgtgtc cgagggtact aaggccatca ccaagtacac cagcgctaag
     3901 gatccccggg taccggtcgc caccatggtg agcaagggcg aggagctgtt caccggggtg
     3961 gtgcccatcc tggtcgagct ggacggcgac gtaaacggcc acaagttcag cgtgtccggc
     4021 gagggcgagg gcgatgccac ctacggcaag ctgaccctga agttcatctg caccaccggc
     4081 aagctgcccg tgccctggcc caccctcgtg accaccttcg gctacggcct gatgtgcttc
     4141 gcccgctacc ccgaccacat gaagcagcac gacttcttca agtccgccat gcccgaaggc
     4201 tacgtccagg agcgcaccat cttcttcaag gacgacggca actacaagac ccgcgccgag
     4261 gtgaagttcg agggcgacac cctggtgaac cgcatcgagc tgaagggcat cgacttcaag
     4321 gaggacggca acatcctggg gcacaagctg gagtacaact acaacagcca caacgtctat
     4381 atcatggccg acaagcagaa gaacggcatc aaggtgaact tcaagatccg ccacaacatc
     4441 gaggacggca gcgtgcagct cgccgaccac taccagcaga acacccccat cggcgacggc
     4501 cccgtgctgc tgcccgacaa ccactacctg agctaccagt ccgccctgag caaagacccc
     4561 aacgagaagc gcgatcacat ggtcctgctg gagttcgtga ccgccgccgg gatcactctc
     4621 ggcatggacg agctgtacaa gtaggtcgac ctcgaggggg ggcccggtac caggtaagtg
     4681 tacccaattc gccctatagt gagtcgtatt acaattcact cgatcgccct tcccaacagt
     4741 tgcgcagcct gaatggcgaa tggagatcca atttttaagt gtataatgtg ttaaactact
     4801 gattctaatt gtttgtgtat tttagattca cagtcccaag gctcatttca ggcccctcag
     4861 tcctcacagt ctgttcatga tcataatcag ccataccaca tttgtagagg ttttacttgc
     4921 tttaaaaaac ctcccacacc tccccctgaa cctgaaacat aaaatgaatg caattgttgt
     4981 tgttaacttg tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt
     5041 cacaaataaa gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt
     5101 atcttaacgc gtaaattgta agcgtctcaa atatgtatcc gctcatgaga caataaccct
     5161 gataaatgct tcaataatat tgaaaaagga agagtatgag tattcaacat ttccgtgtcg
     5221 cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg
     5281 tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt gggttacatc gaactggatc
     5341 tcaacagcgg taagatcctt gagagttttc gccccgaaga acgttttcca atgatgagca
     5401 cttttaaagt tctgctatgt ggcgcggtat tatcccgtat tgacgccggg caagagcaac
     5461 tcggtcgccg catacactat tctcagaatg acttggttga gtactcacca gtcacagaaa
     5521 agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata accatgagtg
     5581 ataacactgc ggccaactta cttctgacaa cgatcggagg accgaaggag ctaaccgctt
     5641 ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg gagctgaatg
     5701 aagccatacc aaacgacgag cgtgacacca cgatgcctgt agcaatggca acaacgttgc
     5761 gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta atagactgga
     5821 tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta
     5881 ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc
     5941 cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg
     6001 atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat tggtaactgt
     6061 cagaccaagt ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa
     6121 ggatctaggt gaagatcctt tttgataatc
//
