LOCUS       sequence-394327-        3484 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  mini circle middle entry plasmid for 2a-mCherry-pA ORF 3 (with ATG) 
            with flanking BsaI cut sites.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3484)
  AUTHORS   Kang Lab
  TITLE     mini-Golden version1
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 3484)
  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            "Unpublished"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 with ATG
FEATURES             Location/Qualifiers
     source          1..3484
                     /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..817
                     /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(252..270)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     520..539
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     polyA_signal    847..981
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(871..890)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     925..944
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             1418..2209
                     /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      2302..2890
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2791..2810
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3044..3061
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(3220..3247)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(3339..3425)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     join(3474..3484,1..12)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
ORIGIN
        1 cgttgtaaaa cgacggccag tcttaagctc gggcccggtc tctagtcccg ctacaaactt
       61 ctctctgctg aagcaggctg gagacgtgga ggagaaccct ggacctatgg tgagcaaggg
      121 cgaggaggat aacatggcca tcatcaagga gttcatgcgc ttcaaggtgc acatggaggg
      181 ctccgtgaac ggccacgagt tcgagatcga gggcgagggc gagggccgcc cctacgaggg
      241 cacccagacc gccaagctga aggtgaccaa gggtggcccc ctgcccttcg cctgggacat
      301 cctgtcccct cagttcatgt acggctccaa ggcctacgtg aagcacccag ccgacatccc
      361 cgactacttg aagctgtcct tccccgaggg cttcaagtgg gagcgcgtga tgaacttcga
      421 ggacggcggc gtggtgaccg tgacccagga ctcctccctg caggacggcg agttcatcta
      481 caaggtgaag ctgcgcggca ccaacttccc ctccgacggc cccgtaatgc agaagaagac
      541 catgggctgg gaggcctcct ccgagcggat gtaccccgag gacggcgccc tgaagggcga
      601 gatcaagcag aggctgaagc tgaaggacgg cggccactac gacgctgagg tcaagaccac
      661 ctacaaggcc aagaagcccg tgcagctgcc cggcgcctac aacgtcaaca tcaagttgga
      721 catcacctcc cacaacgagg actacaccat cgtggaacag tacgaacgcg ccgagggccg
      781 ccactccacc ggcggcatgg acgagctgta caagtaagct agaactatag tgagtcgtat
      841 tacgtagatc cagacatgat aagatacatt gatgagtttg gacaaaccac aactagaatg
      901 cagtgaaaaa aatgctttat ttgtgaaatt tgtgatgcta ttgctttatt tgtaaccatt
      961 ataagctgca ataaacaagt taacaacaac aattgcattc attttatgtt tcaggttcag
     1021 ggggaggtgt gggaggtttt ttaattcctc aagagacctc tagaccagcc aggacagaaa
     1081 tgcctcgact tcgctgctac ccaaggttgc cgggtgacgc acaccgtgga aacggatgaa
     1141 ggcacgaacc cagtggacat aagcctgttc ggttcgtaag ctgtaatgca agtagcgtat
     1201 gcgctcacgc aactggtcca gaaccttgac cgaacgcagc ggtggtaacg gcgcagtggc
     1261 ggttttcatg gcttgttatg actgtttttt tggggtacag tctatgcctc gggcatccaa
     1321 gcagcaagcg cgttacgccg tgggtcgatg tttgatgtta tggagcagca acgatgttac
     1381 gcagcagggc agtcgcccta aaacaaagtt aaacattatg agggaagcgg tgatcgccga
     1441 agtatcgact caactatcag aggtagttgg cgtcatcgag cgccatctcg aaccgacgtt
     1501 gctggccgta catttgtacg gctccgcagt ggatggcggc ctgaagccac acagtgatat
     1561 tgatttgctg gttacggtga ccgtaaggct tgatgaaaca acgcggcgag ctttgatcaa
     1621 cgaccttttg gaaacttcgg cttcccctgg agagagcgag attctccgcg ctgtagaagt
     1681 caccattgtt gtgcacgacg acatcattcc gtggcgttat ccagctaagc gcgaactgca
     1741 atttggagaa tggcagcgca atgacattct tgcaggtatc ttcgagccag ccacgatcga
     1801 cattgatctg gctatcttgc tgacaaaagc aagagaacat agcgttgcct tggtaggtcc
     1861 agcggcggag gaactctttg atccggttcc tgaacaggat ctatttgagg cgctaaatga
     1921 aaccttaacg ctatggaact cgccgcccga ctgggctggc gatgagcgaa atgtagtgct
     1981 tacgttgtcc cgcatttggt acagcgcagt aaccggcaaa atcgcgccga aggatgtcgc
     2041 tgccgactgg gcaatggagc gcctgccggc ccagtatcag cccgtcatac ttgaagctag
     2101 acaggcttat cttggacaag aagaagatcg cttggcctcg cgcgcagatc agttggaaga
     2161 atttgtccac tacgtgaaag gcgagatcac caaggtagtc ggcaaataac cctcgagcca
     2221 cccatgacca aaatccctta acgtgagtta cgcgtcgttc cactgagcgt cagaccccgt
     2281 agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca
     2341 aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct
     2401 ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgttc ttctagtgta
     2461 gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc tcgctctgct
     2521 aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg ggttggactc
     2581 aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt cgtgcacaca
     2641 gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg agctatgaga
     2701 aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg
     2761 aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt atagtcctgt
     2821 cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag gggggcggag
     2881 cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt
     2941 tgctcacatg ttctttcctg cgttatcccc tgattctgtg gataaccgta ttaccgcctt
     3001 tgagtgagct gataccgctc gccgcagccg aacgaccgag cgcagcgagt cagtgagcga
     3061 ggaagcggaa gagcgcccaa tacgcaaacc gcctctcccc gcgcgttggc cgattcatta
     3121 atgcagctgg cacgacaggt ttcccgactg gaaagcgggc agtgagcgca acgcaattaa
     3181 tacgcgtacc gctagccagg aagagtttgt agaaacgcaa aaaggccatc cgtcaggatg
     3241 gccttctgct tagtttgatg cctggcagtt tatggcgggc gtcctgcccg ccaccctccg
     3301 ggccgttgct tcacaacgtt caaatccgct cccggcggat ttgtcctact caggagagcg
     3361 ttcaccgaca aacaacagat aaaacgaaag gcccagtctt ccgactgagc ctttcgtttt
     3421 atttgatgcc tggcagttcc ctactctcgc gttaacgcta gcatggatgt tttcccagtc
     3481 acga
//
