LOCUS       sequence-394268-        3480 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  mini circle middle entry plasmid for 2a-mCherry-pA ORF 2 (without 
            ATG) with flanking BsaI cut sites.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3480)
  AUTHORS   Kang Lab
  TITLE     mini-Golden version1
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 3480)
  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: 
            "Unpublished"
COMMENT     Sequence Label: pMC-ME_2a-mCherry-pA ORF-2 without ATG
FEATURES             Location/Qualifiers
     source          1..3480
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     7..24
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     8..24
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             52..108
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             109..816
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="VSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEGT
                     QTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFE
                     DGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALKG
                     EIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERAE
                     GRHSTGGMDELYK"
     primer_bind     complement(251..269)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     519..538
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     polyA_signal    846..980
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(870..889)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     924..943
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             1417..2208
                     /codon_start=1
                     /gene="aadA"
                     /product="aminoglycoside adenylyltransferase (Murphy, 
                     1985)"
                     /label=SmR
                     /note="confers resistance to spectinomycin and 
                     streptomycin"
                     /translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
                     SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
                     RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
                     EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
                     VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK"
     rep_origin      2301..2889
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2790..2809
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3043..3060
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(3219..3246)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(3338..3424)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     join(3473..3480,1..15)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
ORIGIN
        1 cgacgttgta aaacgacggc cagtcttaag ctcgggcccg gtctctagtc cgctacaaac
       61 ttctctctgc tgaagcaggc tggagacgtg gaggagaacc ctggacctgt gagcaagggc
      121 gaggaggata acatggccat catcaaggag ttcatgcgct tcaaggtgca catggagggc
      181 tccgtgaacg gccacgagtt cgagatcgag ggcgagggcg agggccgccc ctacgagggc
      241 acccagaccg ccaagctgaa ggtgaccaag ggtggccccc tgcccttcgc ctgggacatc
      301 ctgtcccctc agttcatgta cggctccaag gcctacgtga agcacccagc cgacatcccc
      361 gactacttga agctgtcctt ccccgagggc ttcaagtggg agcgcgtgat gaacttcgag
      421 gacggcggcg tggtgaccgt gacccaggac tcctccctgc aggacggcga gttcatctac
      481 aaggtgaagc tgcgcggcac caacttcccc tccgacggcc ccgtaatgca gaagaagacc
      541 atgggctggg aggcctcctc cgagcggatg taccccgagg acggcgccct gaagggcgag
      601 atcaagcaga ggctgaagct gaaggacggc ggccactacg acgctgaggt caagaccacc
      661 tacaaggcca agaagcccgt gcagctgccc ggcgcctaca acgtcaacat caagttggac
      721 atcacctccc acaacgagga ctacaccatc gtggaacagt acgaacgcgc cgagggccgc
      781 cactccaccg gcggcatgga cgagctgtac aagtaagcta gaactatagt gagtcgtatt
      841 acgtagatcc agacatgata agatacattg atgagtttgg acaaaccaca actagaatgc
      901 agtgaaaaaa atgctttatt tgtgaaattt gtgatgctat tgctttattt gtaaccatta
      961 taagctgcaa taaacaagtt aacaacaaca attgcattca ttttatgttt caggttcagg
     1021 gggaggtgtg ggaggttttt taattcctca agagacctct agaccagcca ggacagaaat
     1081 gcctcgactt cgctgctacc caaggttgcc gggtgacgca caccgtggaa acggatgaag
     1141 gcacgaaccc agtggacata agcctgttcg gttcgtaagc tgtaatgcaa gtagcgtatg
     1201 cgctcacgca actggtccag aaccttgacc gaacgcagcg gtggtaacgg cgcagtggcg
     1261 gttttcatgg cttgttatga ctgttttttt ggggtacagt ctatgcctcg ggcatccaag
     1321 cagcaagcgc gttacgccgt gggtcgatgt ttgatgttat ggagcagcaa cgatgttacg
     1381 cagcagggca gtcgccctaa aacaaagtta aacattatga gggaagcggt gatcgccgaa
     1441 gtatcgactc aactatcaga ggtagttggc gtcatcgagc gccatctcga accgacgttg
     1501 ctggccgtac atttgtacgg ctccgcagtg gatggcggcc tgaagccaca cagtgatatt
     1561 gatttgctgg ttacggtgac cgtaaggctt gatgaaacaa cgcggcgagc tttgatcaac
     1621 gaccttttgg aaacttcggc ttcccctgga gagagcgaga ttctccgcgc tgtagaagtc
     1681 accattgttg tgcacgacga catcattccg tggcgttatc cagctaagcg cgaactgcaa
     1741 tttggagaat ggcagcgcaa tgacattctt gcaggtatct tcgagccagc cacgatcgac
     1801 attgatctgg ctatcttgct gacaaaagca agagaacata gcgttgcctt ggtaggtcca
     1861 gcggcggagg aactctttga tccggttcct gaacaggatc tatttgaggc gctaaatgaa
     1921 accttaacgc tatggaactc gccgcccgac tgggctggcg atgagcgaaa tgtagtgctt
     1981 acgttgtccc gcatttggta cagcgcagta accggcaaaa tcgcgccgaa ggatgtcgct
     2041 gccgactggg caatggagcg cctgccggcc cagtatcagc ccgtcatact tgaagctaga
     2101 caggcttatc ttggacaaga agaagatcgc ttggcctcgc gcgcagatca gttggaagaa
     2161 tttgtccact acgtgaaagg cgagatcacc aaggtagtcg gcaaataacc ctcgagccac
     2221 ccatgaccaa aatcccttaa cgtgagttac gcgtcgttcc actgagcgtc agaccccgta
     2281 gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa
     2341 acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt
     2401 tttccgaagg taactggctt cagcagagcg cagataccaa atactgttct tctagtgtag
     2461 ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta
     2521 atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca
     2581 agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag
     2641 cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa
     2701 agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga
     2761 acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc
     2821 gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc
     2881 ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt
     2941 gctcacatgt tctttcctgc gttatcccct gattctgtgg ataaccgtat taccgccttt
     3001 gagtgagctg ataccgctcg ccgcagccga acgaccgagc gcagcgagtc agtgagcgag
     3061 gaagcggaag agcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa
     3121 tgcagctggc acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat
     3181 acgcgtaccg ctagccagga agagtttgta gaaacgcaaa aaggccatcc gtcaggatgg
     3241 ccttctgctt agtttgatgc ctggcagttt atggcgggcg tcctgcccgc caccctccgg
     3301 gccgttgctt cacaacgttc aaatccgctc ccggcggatt tgtcctactc aggagagcgt
     3361 tcaccgacaa acaacagata aaacgaaagg cccagtcttc cgactgagcc tttcgtttta
     3421 tttgatgcct ggcagttccc tactctcgcg ttaacgctag catggatgtt ttcccagtca
//
