LOCUS       sequence-42845-e        6275 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 6275)
  AUTHORS   Larson DR, Zenklusen D, Wu B, Chao JA, Singer RH
  TITLE     Real-time observation of transcription initiation and elongation on 
            an endogenous yeast gene.
  JOURNAL   Science. 2011 Apr 22;332(6028):475-8.
  PUBMED    21512033
REFERENCE   2  (bases 1 to 6275)
  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: "Science."; 
            date: "2011-04-22"; volume: "332(6028)"; pages: "475-8"
COMMENT     Sequence Label: pDZ251 (pKAN 24xPP7SL cyc)
FEATURES             Location/Qualifiers
     source          1..6275
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(1491..1507)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1492..1508)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     terminator      1510..1757
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(1526..1544)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     protein_bind    1808..1841
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     gene            1896..3252
                     /label=kanMX
                     /note="yeast selectable marker conferring kanamycin 
                     resistance (Wach et al., 1994)"
     promoter        1896..2239
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             2240..3049
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin"
                     /translation="MGKEKTHVSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(2307..2326)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     terminator      3055..3252
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     protein_bind    3315..3348
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     primer_bind     complement(3399..3419)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(3399..3417)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3438..3454)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3438..3454)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3451..3473)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3462..3478
                     /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(3486..3492,3493..3510,3511..3516))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3531..3552
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3669..3686)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3840..4428)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3920..3939)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4599..5390,5391..5459))
                     /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     5222..5241
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5460..5564)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      5591..6046
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5678..5697)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5888..5909
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6172..6194
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     6186..6203
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     6187..6203
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6213..6232
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        6213..6231
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     join(6264..6275,1..5)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     join(6264..6275,1..5)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
ORIGIN
        1 ggatcctaag gtacctaatt gcctagaaag gagcagacga tatggcgtcg ctccctgcag
       61 gtcgactcta gaaaccagca gagcatatgg gctcgctggc tgcagtattc ccgggttcat
      121 tagatcctaa ggtacctaat tgcctagaaa ggagcagacg atatggcgtc gctccctgca
      181 ggtcgactct agaaaccagc agagcatatg ggctcgctgg ctgcagtatt cccgggttca
      241 ttagatccta aggtacctaa ttgcctagaa aggagcagac gatatggcgt cgctccctgc
      301 aggtcgactc tagaaaccag cagagcatat gggctcgctg gctgcagtat tcccgggttc
      361 attagatcct aaggtaccta attgcctaga aaggagcaga cgatatggcg tcgctccctg
      421 caggtcgact ctagaaacca gcagagcata tgggctcgct ggctgcagta ttcccgggtt
      481 cattagatcc taaggtacct aattgcctag aaaggagcag acgatatggc gtcgctccct
      541 gcaggtcgac tctagaaacc agcagagcat atgggctcgc tggctgcagt attcccgggt
      601 tcattagatc ctaaggtacc taattgccta gaaaggagca gacgatatgg cgtcgctccc
      661 tgcaggtcga ctctagaaac cagcagagca tatgggctcg ctggctgcag tattcccggg
      721 ttcattagat cctaaggtac ctaattgcct agaaaggagc agacgatatg gcgtcgctcc
      781 ctgcaggtcg actctagaaa ccagcagagc atatgggctc gctggctgca gtattcccgg
      841 gttcattaga tcctaaggta cctaattgcc tagaaaggag cagacgatat ggcgtcgctc
      901 cctgcaggtc gactctagaa accagcagag catatgggct cgctggctgc agtattcccg
      961 ggttcattag atcctaaggt acctaattgc ctagaaagga gcagacgata tggcgtcgct
     1021 ccctgcaggt cgactctaga aaccagcaga gcatatgggc tcgctggctg cagtattccc
     1081 gggttcatta gatcctaagg tacctaattg cctagaaagg agcagacgat atggcgtcgc
     1141 tccctgcagg tcgactctag aaaccagcag agcatatggg ctcgctggct gcagtattcc
     1201 cgggttcatt agatcctaag gtacctaatt gcctagaaag gagcagacga tatggcgtcg
     1261 ctccctgcag gtcgactcta gaaaccagca gagcatatgg gctcgctggc tgcagtattc
     1321 ccgggttcat tagatcctaa ggtacctaat tgcctagaaa ggagcagacg atatggcgtc
     1381 gctccctgca ggtcgactct agaaaccagc agagcatatg ggctcgctgg ctgcagtatt
     1441 cccgggttca ttagatcccc cgggctgcag gaattcgata tcaagcttat cgataccgtc
     1501 gacctcgagt catgtaatta gttatgtcac gcttacattc acgccctccc cccacatccg
     1561 ctctaaccga aaaggaagga gttagacaac ctgaagtcta ggtccctatt tattttttta
     1621 tagttatgtt agtattaaga acgttattta tatttcaaat ttttcttttt tttctgtaca
     1681 gacgcgtgta cgcatgtaac attatactga aaaccttgct tgagaaggtt ttgggacgct
     1741 cgaaggcttt aatttgcggc cggtacccag ctgaagcttc gtacgctgca ggtcgacaac
     1801 ccttaatata acttcgtata atgtatgcta tacgaagtta ttaggtctag agatctgttt
     1861 agcttgcctc gtccccgccg ggtcacccgg ccagcgacat ggaggcccag aataccctcc
     1921 ttgacagtct tgacgtgcgc agctcagggg catgatgtga ctgtcgcccg tacatttagc
     1981 ccatacatcc ccatgtataa tcatttgcat ccatacattt tgatggccgc acggcgcgaa
     2041 gcaaaaatta cggctcctcg ctgcagacct gcgagcaggg aaacgctccc ctcacagacg
     2101 cgttgaattg tccccacgcc gcgcccctgt agagaaatat aaaaggttag gatttgccac
     2161 tgaggttctt ctttcatata cttcctttta aaatcttgct aggatacagt tctcacatca
     2221 catccgaaca taaacaacca tgggtaagga aaagactcac gtttcgaggc cgcgattaaa
     2281 ttccaacatg gatgctgatt tatatgggta taaatgggct cgcgataatg tcgggcaatc
     2341 aggtgcgaca atctatcgat tgtatgggaa gcccgatgcg ccagagttgt ttctgaaaca
     2401 tggcaaaggt agcgttgcca atgatgttac agatgagatg gtcagactaa actggctgac
     2461 ggaatttatg cctcttccga ccatcaagca ttttatccgt actcctgatg atgcatggtt
     2521 actcaccact gcgatccccg gcaaaacagc attccaggta ttagaagaat atcctgattc
     2581 aggtgaaaat attgttgatg cgctggcagt gttcctgcgc cggttgcatt cgattcctgt
     2641 ttgtaattgt ccttttaaca gcgatcgcgt atttcgtctc gctcaggcgc aatcacgaat
     2701 gaataacggt ttggttgatg cgagtgattt tgatgacgag cgtaatggct ggcctgttga
     2761 acaagtctgg aaagaaatgc ataagctttt gccattctca ccggattcag tcgtcactca
     2821 tggtgatttc tcacttgata accttatttt tgacgagggg aaattaatag gttgtattga
     2881 tgttggacga gtcggaatcg cagaccgata ccaggatctt gccatcctat ggaactgcct
     2941 cggtgagttt tctccttcat tacagaaacg gctttttcaa aaatatggta ttgataatcc
     3001 tgatatgaat aaattgcagt ttcatttgat gctcgatgag tttttctaat cagtactgac
     3061 aataaaaaga ttcttgtttt caagaacttg tcatttgtat agttttttta tattgtagtt
     3121 gttctatttt aatcaaatgt tagcgtgatt tatatttttt ttcgcctcga catcatctgc
     3181 ccagatgcga agttaagtgc gcagaaagta atatcatgcg tcaatcgtat gtgaatgctg
     3241 gtcgctatac tgctgtcgat tcgatactaa cgccgccatc cagtgtcgaa aacgagctct
     3301 cgagaaccct taatataact tcgtataatg tatgctatac gaagttatta ggtgatatca
     3361 gatccactag tggcctatgc ggtacccagc ttttgttccc tttagtgagg gttaattgcg
     3421 cgcttggcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc gctcacaatt
     3481 ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta atgagtgagc
     3541 taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc
     3601 cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct
     3661 tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca
     3721 gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac
     3781 atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt
     3841 ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg
     3901 cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc
     3961 tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc
     4021 gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc
     4081 aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac
     4141 tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt
     4201 aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct
     4261 aactacggct acactagaag gacagtattt ggtatctgcg ctctgctgaa gccagttacc
     4321 ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt
     4381 ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg
     4441 atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc
     4501 atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa
     4561 tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag
     4621 gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg
     4681 tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga
     4741 gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag
     4801 cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa
     4861 gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc
     4921 atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca
     4981 aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg
     5041 atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat
     5101 aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc
     5161 aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg
     5221 gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg
     5281 gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt
     5341 gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca
     5401 ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata
     5461 ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac
     5521 atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa
     5581 gtgccacctg acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc
     5641 agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc
     5701 tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg
     5761 ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgattaggg tgatggttca
     5821 cgtagtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc
     5881 tttaatagtg gactcttgtt ccaaactgga acaacactca accctatctc ggtctattct
     5941 tttgatttat aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa
     6001 caaaaattta acgcgaattt taacaaaata ttaacgctta caatttccat tcgccattca
     6061 ggctgcgcaa ctgttgggaa gggcgatcgg tgcgggcctc ttcgctatta cgccagctgg
     6121 cgaaaggggg atgtgctgca aggcgattaa gttgggtaac gccagggttt tcccagtcac
     6181 gacgttgtaa aacgacggcc agtgagcgcg cgtaatacga ctcactatag ggcgaattgg
     6241 agctccaccg cggtggcggc cgctctagaa ctagt
//
