LOCUS       sequence-337490-        4018 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  New measurement standard.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4018)
  AUTHORS   Shao B, Rammohan J, Anderson DA, Alperovich N, Ross D, Voigt CA
  TITLE     Single-cell measurement of plasmid copy number and promoter 
            activity.
  JOURNAL   Nat Commun. 2021 Mar 5;12(1):1475. doi: 10.1038/s41467-021-21734-y.
  PUBMED    33674569
REFERENCE   2  (bases 1 to 4018)
  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: "Nat 
            Commun."; date: "2021-03-5"; volume: "12(1):1475. doi"; pages: " 
            10.1038/s41467-021-21734-y"
COMMENT     Sequence Label: pSB223
FEATURES             Location/Qualifiers
     source          1..4018
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(427..446)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     rep_origin      complement(811..1356)
                     /direction=LEFT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     complement(891..910)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(1445..2260)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(1445..2260)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     2168..2187
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2252..2271
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     promoter        2433..2467
                     /label=BBa_J23101
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.7"
     misc_RNA        2474..2523
                     /label=sTRSV HHRz
                     /note="hammerhead ribozyme from the tobacco ringspot virus 
                     satellite RNA (Khvorova et al., 2003)"
     CDS             join(2573..2575,2576..2578,2579..3292)
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKLHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2618..2639)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2879..2898)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3226..3247
                     /label=EGFP-C
                     /note="EGFP, forward primer"
ORIGIN
        1 atacgaaacg taccgtatcg ttaaggtctg atgatacgaa acgtaccgta tcgttaaggt
       61 aaatgatacg aaacgtaccg tatcgttaag gtatgatcta actactagag tacatgatac
      121 gaaacgtacc gtatcgttaa ggtcatgata cgaaacgtac cgtatcgtta aggttgtatg
      181 atacgaaacg taccgtatcg ttaaggtgca tgatacgaaa cgtaccgtat cgttaaggtc
      241 tgatgatacg aaacgtaccg tatcgttaag gtatgatcta actactagta gcgcccgctg
      301 cagctcggta ccaaattcca gaaaagaggc ctcccgaaag gggggccttt tttcgttttg
      361 gtccgatcct ctacgccgga cgcatcgtgg ccggcatcac cggcgccaca ggtgcggttg
      421 ctggcgccta tatcgccgac atcaccgatg gggaagatcg ggctcgccac ttcgggctca
      481 tgagcaaata ttttatctga ggtgcttcct cgctcactga ctcgctgcac gaggcagacc
      541 tcagcgctag cggagtgtat actggcttac tatgttggca ctgatgaggg tgtcagtgaa
      601 gtgcttcatg tggcaggaga aaaaaggctg caccggtgcg tcagcagaat atgtgataca
      661 ggatatattc cgcttcctcg ctcactgact cgctacgctc ggtcgttcga ctgcggcgag
      721 cggaaatggc ttacgaacgg ggcggagatt tcctggaaga tgccaggaag atacttaaca
      781 gggaagtgag agggccgcgg caaagccgtt tttccatagg ctccgccccc ctgacaagca
      841 tcacgaaatc tgacgctcaa atcagtggtg gcgaaacccg acaggactat aaagatacca
      901 ggcgtttccc cctggcggct ccctcgtgcg ctctcctgtt cctgcctttc ggtttaccgg
      961 tgtcattccg ctgttatggc cgcgtttgtc tcattccacg cctgacactc agttccgggt
     1021 aggcagttcg ctccaagctg gactgtatgc acgaaccccc cgttcagtcc gaccgctgcg
     1081 ccttatccgg taactatcgt cttgagtcca acccggaaag acatgcaaaa gcaccactgg
     1141 cagcagccac tggtaattga tttagaggag ttagtcttga agtcatgcgc cggttaaggc
     1201 taaactgaaa ggacaagttt tggtgactgc gctcctccaa gccagttacc tcggttcaaa
     1261 gagttggtag ctcagagaac cttcgaaaaa ccgccctgca aggcggtttt ttcgttttca
     1321 gagcaagaga ttacgcgcag accaaaacga tctcaagaag atcatcttat taaggggtct
     1381 gacgctcagt ggaacgaaaa atcaatctaa agtatatatg agtaaacttg gtctgacagt
     1441 taccttagaa aaactcatcg agcatcaaat gaaactgcaa tttattcata tcaggattat
     1501 caataccata tttttgaaaa agccgtttct gtaatgaagg agaaaactca ccgaggcagt
     1561 tccataggat ggcaagatcc tggtatcggt ctgcgattcc gactcgtcca acatcaatac
     1621 aacctattaa tttcccctcg tcaaaaataa ggttatcaag tgagaaatca ccatgagtga
     1681 cgactgaatc cggtgagaat ggcaaaagct tatgcatttc tttccagact tgttcaacag
     1741 gccagccatt acgctcgtca tcaaaatcac tcgcatcaac caaaccgtta ttcattcgtg
     1801 attgcgcctg agcgagacga aatacgcgat cgctgttaaa aggacaatta caaacaggaa
     1861 tcgaatgcaa ccggcgcagg aacactgcca gcgcatcaac aatattttca cctgaatcag
     1921 gatattcttc taatacctgg aatgctgttt tcccggggat cgcagtggtg agtaaccatg
     1981 catcatcagg agtacggata aaatgcttga tggtcggaag aggcataaat tccgtcagcc
     2041 agtttagtct gaccatctca tctgtaacat cattggcaac gctacctttg ccatgtttca
     2101 gaaacaactc tggcgcatcg ggcttcccat acaatcgata gattgtcgca cctgattgcc
     2161 cgacattatc gcgagcccat ttatacccat ataaatcagc atccatgttg gaatttaatc
     2221 gcggcctgga gcaagacgtt tcccgttgaa tatggctcat aacacccctt gtattactgt
     2281 ttatgtaagc agacagtttt attgttcatg atgatatatt tttatcttgt gcaatgtaca
     2341 tcagagattt tgagacacaa ccaattattg aaggcctccc taacgggggg cctttttttg
     2401 tttctggtct cccgcttgat aagtccctaa cttttacagc tagctcagtc ctaggtatta
     2461 tgctagcctg aagctgtcac cggatgtgct ttccggtctg atgagtccgt gaggacgaaa
     2521 cagcctctac aaataatttt gtttaatact agagaaagag gggaaatact agatggtgag
     2581 caagggcgag gagctgttca ccggggtggt gcccatcctg gtcgagctgg acggcgacgt
     2641 aaacggccac aagttcagcg tgtccggcga gggcgagggc gatgccacct acggcaagct
     2701 gaccctgaag ttcatctgca ccacaggcaa gctgcccgtg ccctggccca ccctcgtgac
     2761 caccttcggc tacggcctgc aatgcttcgc ccgctacccc gaccacatga agctgcacga
     2821 cttcttcaag tccgccatgc ccgaaggcta cgtccaggag cgcaccatct tcttcaagga
     2881 cgacggcaac tacaagaccc gcgccgaggt gaagttcgag ggcgacaccc tggtgaaccg
     2941 catcgagctg aagggcatcg acttcaagga ggacggcaac atcctggggc acaagctgga
     3001 gtacaactac aacagccaca acgtctatat catggccgac aagcagaaga acggcatcaa
     3061 ggtgaacttc aagatccgcc acaacatcga ggacggcagc gtgcagctcg ccgaccacta
     3121 ccagcagaac accccaatcg gcgacggccc cgtgctgctg cccgacaacc actaccttag
     3181 ctaccagtcc gccctgagca aagaccccaa cgagaagcgc gatcacatgg tcctgctgga
     3241 gttcgtgacc gccgccggga tcactctcgg catggacgag ctgtacaagt aaggatccta
     3301 aggtacctaa ttgcctagaa aggagcagac gatatggcgt cgctccctgc aggtcgactc
     3361 tagaaaccag cagagcatat gggctcgctg gctgcagtat tcccgggttc attagatcct
     3421 aaggtaccta attgcctaga aaggagcaga cgatatggcg tcgctccctg caggtcgact
     3481 ctagaaacca gcagagcata tgggctcgct ggctgcagta ttcccgggtt cattagatcc
     3541 taaggtacct aattgcctag aaaggagcag acgatatggc gtcgctccct gcaggtcgac
     3601 tctagaaacc agcagagcat atgggctcgc tggctgcagt attcccgggt tcattagatc
     3661 tcgcgaaggg cgaattcgcg gccggttcta gagttggtaa cgaatcagac aattgacggc
     3721 tcgagggagt agcatagggt ttgcagaatc cctgcttcgt ccatttgaca ggcacattat
     3781 gcatcgatga taagctgtca aacatgagca tactagagta catgatacga aacgtaccgt
     3841 atcgttaagg tgcatgatac gaaacgtacc gtatcgttaa ggttgtatga tacgaaacgt
     3901 accgtatcgt taaggtgcat gatacgaaac gtaccgtatc gttaaggtct gatgatacga
     3961 aacgtaccgt atcgttaagg taaatgatac gaaacgtacc gtatcgttaa ggtgcatg
//
