LOCUS       sequence-394326-        3481 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  mini circle middle entry plasmid for 2a-mCherry-pA ORF 3 (without 
            ATG) with flanking BsaI cut sites .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3481)
  AUTHORS   Kang Lab
  TITLE     mini-Golden version1
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 3481)
  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            "Unpublished"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-3 without ATG
FEATURES             Location/Qualifiers
     source          1..3481
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     4..21
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     5..21
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             50..106
                     /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             107..814
                     /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(249..267)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     517..536
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     polyA_signal    844..978
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(868..887)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     922..941
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             1415..2206
                     /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      2299..2887
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2788..2807
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3041..3058
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(3217..3244)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(3336..3422)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     join(3471..3481,1..12)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
ORIGIN
        1 cgttgtaaaa cgacggccag tcttaagctc gggcccggtc tctagtcccg ctacaaactt
       61 ctctctgctg aagcaggctg gagacgtgga ggagaaccct ggacctgtga gcaagggcga
      121 ggaggataac atggccatca tcaaggagtt catgcgcttc aaggtgcaca tggagggctc
      181 cgtgaacggc cacgagttcg agatcgaggg cgagggcgag ggccgcccct acgagggcac
      241 ccagaccgcc aagctgaagg tgaccaaggg tggccccctg cccttcgcct gggacatcct
      301 gtcccctcag ttcatgtacg gctccaaggc ctacgtgaag cacccagccg acatccccga
      361 ctacttgaag ctgtccttcc ccgagggctt caagtgggag cgcgtgatga acttcgagga
      421 cggcggcgtg gtgaccgtga cccaggactc ctccctgcag gacggcgagt tcatctacaa
      481 ggtgaagctg cgcggcacca acttcccctc cgacggcccc gtaatgcaga agaagaccat
      541 gggctgggag gcctcctccg agcggatgta ccccgaggac ggcgccctga agggcgagat
      601 caagcagagg ctgaagctga aggacggcgg ccactacgac gctgaggtca agaccaccta
      661 caaggccaag aagcccgtgc agctgcccgg cgcctacaac gtcaacatca agttggacat
      721 cacctcccac aacgaggact acaccatcgt ggaacagtac gaacgcgccg agggccgcca
      781 ctccaccggc ggcatggacg agctgtacaa gtaagctaga actatagtga gtcgtattac
      841 gtagatccag acatgataag atacattgat gagtttggac aaaccacaac tagaatgcag
      901 tgaaaaaaat gctttatttg tgaaatttgt gatgctattg ctttatttgt aaccattata
      961 agctgcaata aacaagttaa caacaacaat tgcattcatt ttatgtttca ggttcagggg
     1021 gaggtgtggg aggtttttta attcctcaag agacctctag accagccagg acagaaatgc
     1081 ctcgacttcg ctgctaccca aggttgccgg gtgacgcaca ccgtggaaac ggatgaaggc
     1141 acgaacccag tggacataag cctgttcggt tcgtaagctg taatgcaagt agcgtatgcg
     1201 ctcacgcaac tggtccagaa ccttgaccga acgcagcggt ggtaacggcg cagtggcggt
     1261 tttcatggct tgttatgact gtttttttgg ggtacagtct atgcctcggg catccaagca
     1321 gcaagcgcgt tacgccgtgg gtcgatgttt gatgttatgg agcagcaacg atgttacgca
     1381 gcagggcagt cgccctaaaa caaagttaaa cattatgagg gaagcggtga tcgccgaagt
     1441 atcgactcaa ctatcagagg tagttggcgt catcgagcgc catctcgaac cgacgttgct
     1501 ggccgtacat ttgtacggct ccgcagtgga tggcggcctg aagccacaca gtgatattga
     1561 tttgctggtt acggtgaccg taaggcttga tgaaacaacg cggcgagctt tgatcaacga
     1621 ccttttggaa acttcggctt cccctggaga gagcgagatt ctccgcgctg tagaagtcac
     1681 cattgttgtg cacgacgaca tcattccgtg gcgttatcca gctaagcgcg aactgcaatt
     1741 tggagaatgg cagcgcaatg acattcttgc aggtatcttc gagccagcca cgatcgacat
     1801 tgatctggct atcttgctga caaaagcaag agaacatagc gttgccttgg taggtccagc
     1861 ggcggaggaa ctctttgatc cggttcctga acaggatcta tttgaggcgc taaatgaaac
     1921 cttaacgcta tggaactcgc cgcccgactg ggctggcgat gagcgaaatg tagtgcttac
     1981 gttgtcccgc atttggtaca gcgcagtaac cggcaaaatc gcgccgaagg atgtcgctgc
     2041 cgactgggca atggagcgcc tgccggccca gtatcagccc gtcatacttg aagctagaca
     2101 ggcttatctt ggacaagaag aagatcgctt ggcctcgcgc gcagatcagt tggaagaatt
     2161 tgtccactac gtgaaaggcg agatcaccaa ggtagtcggc aaataaccct cgagccaccc
     2221 atgaccaaaa tcccttaacg tgagttacgc gtcgttccac tgagcgtcag accccgtaga
     2281 aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac
     2341 aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt
     2401 tccgaaggta actggcttca gcagagcgca gataccaaat actgttcttc tagtgtagcc
     2461 gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat
     2521 cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag
     2581 acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc
     2641 cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag
     2701 cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac
     2761 aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg
     2821 gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct
     2881 atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc
     2941 tcacatgttc tttcctgcgt tatcccctga ttctgtggat aaccgtatta ccgcctttga
     3001 gtgagctgat accgctcgcc gcagccgaac gaccgagcgc agcgagtcag tgagcgagga
     3061 agcggaagag cgcccaatac gcaaaccgcc tctccccgcg cgttggccga ttcattaatg
     3121 cagctggcac gacaggtttc ccgactggaa agcgggcagt gagcgcaacg caattaatac
     3181 gcgtaccgct agccaggaag agtttgtaga aacgcaaaaa ggccatccgt caggatggcc
     3241 ttctgcttag tttgatgcct ggcagtttat ggcgggcgtc ctgcccgcca ccctccgggc
     3301 cgttgcttca caacgttcaa atccgctccc ggcggatttg tcctactcag gagagcgttc
     3361 accgacaaac aacagataaa acgaaaggcc cagtcttccg actgagcctt tcgttttatt
     3421 tgatgcctgg cagttcccta ctctcgcgtt aacgctagca tggatgtttt cccagtcacg
     3481 a
//
