LOCUS       sequence-394271-        3483 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  mini circle middle entry plasmid for 2a-mCherry-pA ORF 2 (with ATG) 
            with flanking BsaI cut sites.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3483)
  AUTHORS   Kang Lab
  TITLE     mini-Golden version1
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 3483)
  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            "Unpublished"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 with ATG
FEATURES             Location/Qualifiers
     source          1..3483
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1..18
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     2..18
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             46..102
                     /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             103..813
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(248..266)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     516..535
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     polyA_signal    843..977
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(867..886)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     921..940
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             1414..2205
                     /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      2298..2886
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2787..2806
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3040..3057
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(3216..3243)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(3335..3421)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     join(3470..3483,1..9)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
ORIGIN
        1 tgtaaaacga cggccagtct taagctcggg cccggtctct agtccgctac aaacttctct
       61 ctgctgaagc aggctggaga cgtggaggag aaccctggac ctatggtgag caagggcgag
      121 gaggataaca tggccatcat caaggagttc atgcgcttca aggtgcacat ggagggctcc
      181 gtgaacggcc acgagttcga gatcgagggc gagggcgagg gccgccccta cgagggcacc
      241 cagaccgcca agctgaaggt gaccaagggt ggccccctgc ccttcgcctg ggacatcctg
      301 tcccctcagt tcatgtacgg ctccaaggcc tacgtgaagc acccagccga catccccgac
      361 tacttgaagc tgtccttccc cgagggcttc aagtgggagc gcgtgatgaa cttcgaggac
      421 ggcggcgtgg tgaccgtgac ccaggactcc tccctgcagg acggcgagtt catctacaag
      481 gtgaagctgc gcggcaccaa cttcccctcc gacggccccg taatgcagaa gaagaccatg
      541 ggctgggagg cctcctccga gcggatgtac cccgaggacg gcgccctgaa gggcgagatc
      601 aagcagaggc tgaagctgaa ggacggcggc cactacgacg ctgaggtcaa gaccacctac
      661 aaggccaaga agcccgtgca gctgcccggc gcctacaacg tcaacatcaa gttggacatc
      721 acctcccaca acgaggacta caccatcgtg gaacagtacg aacgcgccga gggccgccac
      781 tccaccggcg gcatggacga gctgtacaag taagctagaa ctatagtgag tcgtattacg
      841 tagatccaga catgataaga tacattgatg agtttggaca aaccacaact agaatgcagt
      901 gaaaaaaatg ctttatttgt gaaatttgtg atgctattgc tttatttgta accattataa
      961 gctgcaataa acaagttaac aacaacaatt gcattcattt tatgtttcag gttcaggggg
     1021 aggtgtggga ggttttttaa ttcctcaaga gacctctaga ccagccagga cagaaatgcc
     1081 tcgacttcgc tgctacccaa ggttgccggg tgacgcacac cgtggaaacg gatgaaggca
     1141 cgaacccagt ggacataagc ctgttcggtt cgtaagctgt aatgcaagta gcgtatgcgc
     1201 tcacgcaact ggtccagaac cttgaccgaa cgcagcggtg gtaacggcgc agtggcggtt
     1261 ttcatggctt gttatgactg tttttttggg gtacagtcta tgcctcgggc atccaagcag
     1321 caagcgcgtt acgccgtggg tcgatgtttg atgttatgga gcagcaacga tgttacgcag
     1381 cagggcagtc gccctaaaac aaagttaaac attatgaggg aagcggtgat cgccgaagta
     1441 tcgactcaac tatcagaggt agttggcgtc atcgagcgcc atctcgaacc gacgttgctg
     1501 gccgtacatt tgtacggctc cgcagtggat ggcggcctga agccacacag tgatattgat
     1561 ttgctggtta cggtgaccgt aaggcttgat gaaacaacgc ggcgagcttt gatcaacgac
     1621 cttttggaaa cttcggcttc ccctggagag agcgagattc tccgcgctgt agaagtcacc
     1681 attgttgtgc acgacgacat cattccgtgg cgttatccag ctaagcgcga actgcaattt
     1741 ggagaatggc agcgcaatga cattcttgca ggtatcttcg agccagccac gatcgacatt
     1801 gatctggcta tcttgctgac aaaagcaaga gaacatagcg ttgccttggt aggtccagcg
     1861 gcggaggaac tctttgatcc ggttcctgaa caggatctat ttgaggcgct aaatgaaacc
     1921 ttaacgctat ggaactcgcc gcccgactgg gctggcgatg agcgaaatgt agtgcttacg
     1981 ttgtcccgca tttggtacag cgcagtaacc ggcaaaatcg cgccgaagga tgtcgctgcc
     2041 gactgggcaa tggagcgcct gccggcccag tatcagcccg tcatacttga agctagacag
     2101 gcttatcttg gacaagaaga agatcgcttg gcctcgcgcg cagatcagtt ggaagaattt
     2161 gtccactacg tgaaaggcga gatcaccaag gtagtcggca aataaccctc gagccaccca
     2221 tgaccaaaat cccttaacgt gagttacgcg tcgttccact gagcgtcaga ccccgtagaa
     2281 aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca
     2341 aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt
     2401 ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgttcttct agtgtagccg
     2461 tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc
     2521 ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga
     2581 cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc
     2641 agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc
     2701 gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca
     2761 ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg
     2821 tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta
     2881 tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct
     2941 cacatgttct ttcctgcgtt atcccctgat tctgtggata accgtattac cgcctttgag
     3001 tgagctgata ccgctcgccg cagccgaacg accgagcgca gcgagtcagt gagcgaggaa
     3061 gcggaagagc gcccaatacg caaaccgcct ctccccgcgc gttggccgat tcattaatgc
     3121 agctggcacg acaggtttcc cgactggaaa gcgggcagtg agcgcaacgc aattaatacg
     3181 cgtaccgcta gccaggaaga gtttgtagaa acgcaaaaag gccatccgtc aggatggcct
     3241 tctgcttagt ttgatgcctg gcagtttatg gcgggcgtcc tgcccgccac cctccgggcc
     3301 gttgcttcac aacgttcaaa tccgctcccg gcggatttgt cctactcagg agagcgttca
     3361 ccgacaaaca acagataaaa cgaaaggccc agtcttccga ctgagccttt cgttttattt
     3421 gatgcctggc agttccctac tctcgcgtta acgctagcat ggatgttttc ccagtcacga
     3481 cgt
//
