LOCUS       sequence-56686-e        4468 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4468)
  AUTHORS   Phong C, Markson JS, Wilhoite CM, Rust MJ
  TITLE     Robust and tunable circadian rhythms from differentially sensitive 
            catalytic domains.
  JOURNAL   Proc Natl Acad Sci U S A. 2013 Jan 15;110(3):1124-9. doi: 
            10.1073/pnas.1212113110. Epub 2012 Dec 31.
  PUBMED    23277568
REFERENCE   2  (bases 1 to 4468)
  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"; doi: 
            "10.1073/pnas.1212113110"; journalName: "Proc Natl Acad Sci U S A"; 
            date: "2013-01-15- 15"; volume: "110"; issue: "3"; pages: "1124-9"
COMMENT     Sequence Label: KaiC
FEATURES             Location/Qualifiers
     source          1..4468
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     20..39
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        20..38
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             70..92
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             112..129
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             133..165
                     /codon_start=1
                     /product="leader peptide from bacteriophage T7 gene 10"
                     /label=T7 tag (gene 10 leader)
                     /note="promotes efficient translation in E. coli"
                     /translation="MASMTGGQQMG"
     CDS             join(169..177,178..192)
                     /codon_start=1
                     /product="Xpress(TM) epitope tag, including an enterokinase
                     recognition and cleavage site"
                     /label=Xpress(TM) tag
                     /translation="DLYDDDDK"
     CDS             202..225
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     primer_bind     complement(1869..1887)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1883..1930
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      2027..2482
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2114..2133)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2324..2345
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        2508..2612
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2613..2681,2682..3473)
                     /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(2831..2850)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3644..4232
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4133..4152
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4386..4403
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 gatctcgatc ccgcgaaatt aatacgactc actataggga gaccacaacg gtttccctct
       61 agaaataatt ttgtttaact ttaagaagga gatatacata tgcggggttc tcatcatcat
      121 catcatcatg gtatggctag catgactggt ggacagcaaa tgggtcggga tctgtacgac
      181 gatgacgata aggatccggg actggaagtt ctgttccagg ggcccatgac ttccgctgag
      241 atgactagcc ctaataataa ttctgagcac caagccatcg ctaagatgcg cacgatgatt
      301 gaaggctttg atgatattag tcatggcggt cttccaatcg ggcgatcgac cctcgttagt
      361 ggtacttcag gaaccggcaa gacccttttt tctattcaat ttctctataa cggtattatc
      421 gagtttgatg agcctggggt tttcgttact ttcgaagaaa ccccgcaaga tatcattaaa
      481 aacgcccgta gttttggctg ggatttagcc aagctggtcg atgagggcaa actatttatt
      541 cttgatgctt cacccgatcc agaaggtcaa gaggttgttg gcggcttcga tctctctgct
      601 ctgattgagc ggattaatta tgcaattcaa aagtatcgag cgcggcgggt ttcaattgac
      661 tcggtcacgt ccgttttcca gcaatatgat gcctcttctg tggttcgccg cgaactcttt
      721 cggttggtag ctcgcctaaa acaaattggg gcaactacgg tcatgaccac cgagcgtatc
      781 gaggaatatg gcccgatcgc tcgttacggt gttgaggaat ttgtctccga taacgtcgtg
      841 attctccgca acgttttgga aggggagcgc cgtcgccgca ccctcgaaat cctcaagcta
      901 cgtggcacca gccacatgaa aggggaatat ccgttcacga ttacggatca tggcatcaat
      961 atcttcccgc tcggggcaat gcgccttacg cagcgatcgt cgaacgtgcg tgtttcatct
     1021 ggtgtcgtcc gactcgatga aatgtgtggt gggggcttct ttaaggactc aatcattctg
     1081 gcaactggcg ctacaggcac tggtaaaact ctgttagtta gccgtttcgt tgagaatgct
     1141 tgtgctaaca aagagcgggc gattctgttc gcttatgaag agtcacgagc tcagctgctc
     1201 cgcaatgcct attcatgggg aatggacttt gaggagatgg agcgccaaaa cctcctcaaa
     1261 attgtttgcg cctatcctga atctgcaggt cttgaagacc atttgcagat tattaagtcg
     1321 gagatcaatg actttaagcc agctcgtatt gcaatcgact ccctctctgc tttggcgcgg
     1381 ggcgttagca acaatgcctt ccgccaattt gtaattggtg tcactggcta cgcgaaacaa
     1441 gaagaaatca cgggactatt cacaaatacc agtgatcaat ttatgggagc gcattcgatt
     1501 actgactccc atatctcaac aattacggat acgattatct tgctccaata cgtcgagatt
     1561 cgtggcgaaa tgtcccgcgc cattaacgtc ttcaagatgc gcggatcttg gcatgacaaa
     1621 gcaatccgcg aattcatgat cagcgacaaa gggccggaca tcaaggattc tttccggaac
     1681 tttgagcgga ttatttcagg ttcgccaaca cggattaccg tcgatgagaa aagcgaactc
     1741 tcgcgaattg tgcgcggcgt tcaagaaaaa gggccggaga gctaaagatc tgcagctggt
     1801 accatggaat tcgaagcttg atccggctgc taacaaagcc cgaaaggaag ctgagttggc
     1861 tgctgccacc gctgagcaat aactagcata accccttggg gcctctaaac gggtcttgag
     1921 gggttttttg ctgaaaggag gaactatatc cggatctggc gtaatagcga agaggcccgc
     1981 accgatcgcc cttcccaaca gttgcgcagc ctgaatggcg aatgggacgc gccctgtagc
     2041 ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc
     2101 gccctagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt
     2161 ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac
     2221 ctcgacccca aaaaacttga ttagggtgat ggttcacgta gtgggccatc gccctgatag
     2281 acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa
     2341 actggaacaa cactcaaccc tatctcggtc tattcttttg atttataagg gattttgccg
     2401 atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattttaac
     2461 aaaatattaa cgcttacaat ttaggtggca cttttcgggg aaatgtgcgc ggaaccccta
     2521 tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa taaccctgat
     2581 aaatgcttca ataatattga aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc
     2641 ttattccctt ttttgcggca ttttgccttc ctgtttttgc tcacccagaa acgctggtga
     2701 aagtaaaaga tgctgaagat cagttgggtg cacgagtggg ttacatcgaa ctggatctca
     2761 acagcggtaa gatccttgag agttttcgcc ccgaagaacg ttttccaatg atgagcactt
     2821 ttaaagttct gctatgtggc gcggtattat cccgtattga cgccgggcaa gagcaactcg
     2881 gtcgccgcat acactattct cagaatgact tggttgagta ctcaccagtc acagaaaagc
     2941 atcttacgga tggcatgaca gtaagagaat tatgcagtgc tgccataacc atgagtgata
     3001 acactgcggc caacttactt ctgacaacga tcggaggacc gaaggagcta accgcttttt
     3061 tgcacaacat gggggatcat gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag
     3121 ccataccaaa cgacgagcgt gacaccacga tgcctgtagc aatggcaaca acgttgcgca
     3181 aactattaac tggcgaacta cttactctag cttcccggca acaattaata gactggatgg
     3241 aggcggataa agttgcagga ccacttctgc gctcggccct tccggctggc tggtttattg
     3301 ctgataaatc tggagccggt gagcgtgggt ctcgcggtat cattgcagca ctggggccag
     3361 atggtaagcc ctcccgtatc gtagttatct acacgacggg gagtcaggca actatggatg
     3421 aacgaaatag acagatcgct gagataggtg cctcactgat taagcattgg taactgtcag
     3481 accaagttta ctcatatata ctttagattg atttaaaact tcatttttaa tttaaaagga
     3541 tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt gagttttcgt
     3601 tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc
     3661 tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc
     3721 cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac
     3781 caaatactgt tcttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac
     3841 cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt
     3901 cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct
     3961 gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat
     4021 acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt
     4081 atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg
     4141 cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt
     4201 gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt
     4261 tcctggcctt ttgctggcct tttgctcaca tgttctttcc tgcgttatcc cctgattctg
     4321 tggataaccg tattaccgcc tttgagtgag ctgataccgc tcgccgcagc cgaacgaccg
     4381 agcgcagcga gtcagtgagc gaggaagcgg aagagcgccc aatacgcaaa ccgcctctcc
     4441 ccgcgcgttg gccgattcat taatgcag
//
