LOCUS       sequence-370057-        6711 bp    DNA     circular SYN 14-APR-2025
DEFINITION  Expresses FLAG-BMAL1-S519A in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6711)
  AUTHORS   Li H, Ou J, Li Y, Xu N, Li Q, Wu P, Peng C, Tang YC, Chang HC
  TITLE     ISX-9 potentiates CaMKIIdelta-mediated BMAL1 activation to enhance 
            circadian amplitude.
  JOURNAL   Commun Biol. 2022 Jul 28;5(1):750. doi: 10.1038/s42003-022-03725-x.
  PUBMED    35902736
REFERENCE   2  (bases 1 to 6711)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Commun 
            Biol."; date: "2022-07-28"; volume: "5(1):750. doi"; pages: " 
            10.1038/s42003-022-03725-x"
COMMENT     Sequence Label: pCAGGS-FLAG-BMAL1-S519A
FEATURES             Location/Qualifiers
     source          1..6711
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        5..384
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        386..661
                     /label=chicken beta-actin promoter
     intron          662..1678
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1602..1624
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1686..1705
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     CDS             1739..1762
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     primer_bind     complement(3733..3752)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    3798..3853
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(3852..3871)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(4214..4230)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4214..4230)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4227..4249)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4238..4254
                     /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(join(4262..4268,4269..4286,4287..4292))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4307..4328
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        4387..4582
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      4433..4568
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    4588..4722
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4625..4644)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4679..4698
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4790..4807)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4961..5549)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5041..5060)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5720..6511,6512..6580))
                     /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     6343..6362
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6581..6685)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ggtcgacatt gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat
       61 agcccatata tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg
      121 cccaacgacc cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata
      181 gggactttcc attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta
      241 catcaagtgt atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc
      301 gcctggcatt atgcccagta catgacctta tgggactttc ctacttggca gtacatctac
      361 gtattagtca tcgctattac catggtcgag gtgagcccca cgttctgctt cactctcccc
      421 atctcccccc cctccccacc cccaattttg tatttattta ttttttaatt attttgtgca
      481 gcgatggggg cggggggggg gggggggcgc gccrggsggg gsggggsggg gsgrggggsg
      541 gggsggggsg aggcggagag gtgcggcggc agccaatcag agcggcgcgc tccgaaagtt
      601 tccttttatg gcgaggcggc ggcggcggcg gccctataaa aagcgaagcg cgcggcgggc
      661 gggagtcgct gcgcgctgcc ttcgccccgt gccccgctcc gccgccgcct cgcgccgccc
      721 gccccggctc tgactgaccg cgttactccc acaggtgagc gggcgggacg gcccttctcc
      781 tccgggctgt aattagcgct tggtttaatg acggcttgtt tcttttctgt ggctgcgtga
      841 aagccttgag gggctccggg agggcccttt gtgcgggggg agcggctcgg ggggtgcgtg
      901 cgtgtgtgtg tgcgtgggga gcgccgcgtg cggctccgcg ctgcccggcg gctgtgagcg
      961 ctgcgggcgc ggcgcggggc tttgtgcgct ccgcagtgtg cgcgagggga gcgcggccgg
     1021 gggcggtgcc ccgcggtgcg gggggggctg cgaggggaac aaaggctgcg tgcggggtgt
     1081 gtgcgtgggg gggtgagcag ggggtgtggg cgcgtcggtc gggctgcaac cccccctgca
     1141 cccccctccc cgagttgctg agcacggccc ggcttcgggt gcggggctcc gtacggggcg
     1201 tggcgcgggg ctcgccgtgc cgggcggggg gtggcggcag gtgggggtgc cgggcggggc
     1261 ggggccgcct cgggccgggg agggctcggg ggaggggcgc ggcggccccc ggagcgccgg
     1321 cggctgtcga ggcgcggcga gccgcagcca ttgcctttta tggtaatcgt gcgagagggc
     1381 gcagggactt cctttgtccc aaatctgtgc ggagccgaaa tctgggaggc gccgccgcac
     1441 cccctctagc gggcgcgggg cgaagcggtg cggcgccggc aggaaggaaa tgggcgggga
     1501 gggccttcgt gcgtcgccgc gccgccgtcc ccttctccct ctccagcctc ggggctgtcc
     1561 gcggggggac ggctgccttc gggggggacg gggcagggcg gggttcggct tctggcgtgt
     1621 gaccggcggc tctagagcct ctgctaacca tgttcatgcc ttcttctttt tcctacagct
     1681 cctgggcaac gtgctggtta ttgtgctgtc tcatcatttt ggcaaagaat tcaccatgga
     1741 ttacaaggat gacgacgata agtgcggccg cgcggaccag agaatggaca tttcctcaac
     1801 catcagcgac ttcatgtctc cgggccccac cgacctactc tccggttccc tgggcaccag
     1861 tggtgtggac tgcaatcgca agaggaaagg cagtgccact gactaccaag aaagtatgga
     1921 cacagacaaa gatgaccctc atggaaggtt agaatatgca gaacaccaag gaaggatcaa
     1981 gaatgcaagg gaggcccaca gtcagattga aaagaggcgt cgggacaaaa tgaacagttt
     2041 cattgatgaa ttggcttctt tggtaccaac atgcaatgca atgtccagga agttagataa
     2101 actcaccgtg ctaaggatgg ctgttcagca catgaaaact ttgagaggtg ccaccaaccc
     2161 atacacagaa gcaaactaca agccaacatt tctatcagat gacgaactga aacacctaat
     2221 tctcagggca gcagatggat ttttgtttgt cgtaggatgt gaccgaggga agatcctctt
     2281 tgtctccgag tctgtcttca agatcctcaa ttatagccag aatgacctta ttggccagag
     2341 cttgtttgac tacctgcatc caaaagatat tgccaaagtt aaggaacagc tatcttcctc
     2401 ggacactgcg ccccgggagc gactcattga tgccaagact ggacttccgg ttaaaacgga
     2461 tataacccct gggccctccc ggctatgctc tggagcccgc cgctctttct tctgtagaat
     2521 gaagtgcaac aggccttcag taaaggtgga agataaggac ttcgcctcta cctgttcaaa
     2581 gaaaaaagca gatcgaaaaa gcttctgcac aatccacagc acaggctatt tgaaaagctg
     2641 gccacccacg aagatggggc tggacgaaga caatgagcca gacaacgagg gctgcaacct
     2701 cagctgcctc gttgcaatcg ggcgcctgca ctcgcacatg gttccacaac cagcgaacgg
     2761 ggaaatacgg gtgaaatcta tggagtacgt ttctcgacac gcaatagatg ggaaatttgt
     2821 ttttgtagat cagagggcga cagctatttt ggcgtatcta ccacaggaac ttctaggtac
     2881 atcatgttat gagtattttc atcaagacga cataggacac ctcgcagaat gtcacaggca
     2941 agttttacag acaagagaaa agatcacaac taattgctat aagtttaaga tcaaagatgg
     3001 ttcttttatc acgctacgaa gtcgatggtt cagtttcatg aacccgtgga ccaaggaagt
     3061 tgaatacatt gtctcaacca acactgttgt tttagccaat gtcctggaag gcggggaccc
     3121 aaccttcccg cagctaacag caccccccca cagcatggac agcatgctgc cctctggaga
     3181 aggtggccca aagaggactc atcccactgt cccaggcatt ccagggggaa ccagagccgg
     3241 agcaggaaaa ataggtcgaa tgatcgcgga ggaaatcatg gaaatccaca ggataagagg
     3301 gtcatcgcct tccagctgtg gcgccagccc gctgaacatc acaagtacgc ctccccctga
     3361 tgcctcttct ccaggaggca agaagattct aaatggaggg actccagaca ttccttccac
     3421 tggactatta ccagggcagg ctcaggagac cccagggtat ccctattctg atagttcttc
     3481 tattcttggt gagaaccccc acataggcat cgatatgata gataacgacc aaggatcaag
     3541 tagtcccagt aacgatgagg cagcaatggc tgtcatcatg agcctcttgg aagcagatgc
     3601 ggggctgggt ggccccgttg actttagtga cttgccatgg ccgctgtagg aattcactcc
     3661 tcaggtgcag gctgcctatc agaaggtggt ggctggtgtg gccaatgccc tggctcacaa
     3721 ataccactga gatctttttc cctctgccaa aaattatggg gacatcatga agccccttga
     3781 gcatctgact tctggctaat aaaggaaatt tattttcatt gcaatagtgt gttggaattt
     3841 tttgtgtctc tcactcggaa ggacatatgg gagggcaaat catttaaaac atcagaatga
     3901 gtatttggtt tagagtttgg caacatatgc ccatatgctg gctgccatga acaaaggttg
     3961 gctataaaga ggtcatcagt atatgaaaca gccccctgct gtccattcct tattccatag
     4021 aaaagccttg acttgaggtt agattttttt tatattttgt tttgtgttat ttttttcttt
     4081 aacatcccta aaattttcct tacatgtttt actagccaga tttttcctcc tctcctgact
     4141 actcccagtc atagctgtcc ctcttctctt atggagatcc ctcgacctgc agcccaagct
     4201 tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac
     4261 acaacatacg agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac
     4321 tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc
     4381 ggatccgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc ccatcccgcc
     4441 cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt tttttattta
     4501 tgcagaggcc gaggccgcct cggcctctga gctattccag aagtagtgag gaggcttttt
     4561 tggaggccta ggcttttgca aaaagctaac ttgtttattg cagcttataa tggttacaaa
     4621 taaagcaata gcatcacaaa tttcacaaat aaagcatttt tttcactgca ttctagttgt
     4681 ggtttgtcca aactcatcaa tgtatcttat catgtctgga tccgctgcat taatgaatcg
     4741 gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg
     4801 actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa
     4861 tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc
     4921 aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc
     4981 ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat
     5041 aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc
     5101 cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct
     5161 cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg
     5221 aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc
     5281 cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga
     5341 ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa
     5401 gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta
     5461 gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc
     5521 agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg
     5581 acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga
     5641 tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg
     5701 agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct
     5761 gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg
     5821 agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc
     5881 cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa
     5941 ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc
     6001 cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt
     6061 cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc
     6121 ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt
     6181 tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc
     6241 catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt
     6301 gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata
     6361 gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga
     6421 tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag
     6481 catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa
     6541 aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt
     6601 attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga
     6661 aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct g
//
