LOCUS       sequence-482179-        3642 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Middle entry vector for mini-Golden subcloning platform; mScarlet.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3642)
  TITLE     Mini-Golden Subcloning Platform
REFERENCE   2  (bases 1 to 3642)
  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"
COMMENT     Sequence Label: SA-linker-P2A-mem-mScarlet-ORF-2
FEATURES             Location/Qualifiers
     source          1..3642
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      complement(1..87)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     136..158
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     150..167
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     151..167
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             264..320
                     /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             432..1118
                     /codon_start=1
                     /product="bright cysteine-free monomeric red fluorescent 
                     protein with fast and complete maturation (Gadella et al., 
                     2023)"
                     /label=mScarlet3
                     /translation="MDSTEAVIKEFMRFKVHMEGSMNGHEFEIEGEGEGRPYEGTQTAK
                     LRVTKGGPLPFSWDILSPQFMYGSRAFTKHPADIPDYWKQSFPEGFKWERVMNFEDGGA
                     VSVAQDTSLEDGTLIYKVKLRGTNFPPDGPVMQKKTMGWEASTERLYPEDVVLKGDIKM
                     ALRLKDGGRYLADFKTTYRAKKPVQMPGAFNIDRKLDITSHNEDYTVVEQYERSVARHS
                     TGGSGGS"
     primer_bind     830..849
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     polyA_signal    1151..1285
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1175..1194)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1229..1248
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             1722..2513
                     /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      2606..3194
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3095..3114
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3348..3365
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(3524..3551)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
ORIGIN
        1 atttgtccta ctcaggagag cgttcaccga caaacaacag ataaaacgaa aggcccagtc
       61 ttccgactga gcctttcgtt ttatttgatg cctggcagtt ccctactctc gcgttaacgc
      121 tagcatggat gttttcccag tcacgacgtt gtaaaacgac ggccagtctt aagctcgggc
      181 ccggtctctt cgccactaat ccggagtgtg tgtgtgtttc tgacgtgtct ctctcttccc
      241 tttttttttt cagcggctct ggggctacaa acttctctct gctgaaacaa gctggcgacg
      301 tggaggagaa ccctggacct atgctgtgct gcatcagaag aactaaaccg gttgagaaga
      361 atgaagaggc cgatcaggag ctgcagtcga cggtaccgcg ggcccgggac ccaccggtcg
      421 ccaccaagct tatggatagc accgaggcag tgatcaagga gttcatgcgg ttcaaggtgc
      481 acatggaggg ctccatgaac ggccacgagt tcgagatcga gggcgagggc gagggccgcc
      541 cctacgaggg cacccagacc gccaagctga gggtgaccaa gggtggcccc ctgcccttct
      601 cctgggacat cctgtcccct cagttcatgt acggctccag ggccttcacg aagcaccccg
      661 ccgacatccc cgactactgg aagcagtcct tccccgaggg cttcaagtgg gagcgcgtga
      721 tgaacttcga ggacggcggc gccgtgtccg tggcccagga cacctccctg gaggacggca
      781 ccctgatcta caaggtgaag ctccgcggca ccaacttccc tcctgacggc cccgtaatgc
      841 agaagaagac aatgggctgg gaagcatcca ccgagcggtt gtaccccgag gacgtcgtgc
      901 tgaagggcga cattaagatg gccctgcgcc tgaaggacgg cggccgctac ctggcggact
      961 tcaagaccac ctacagggcc aagaagcccg tgcagatgcc cggcgccttc aacatcgacc
     1021 gcaagttgga catcacatcc cacaacgagg actacaccgt ggtggaacag tacgaacgct
     1081 ccgtggcccg ccactccacc ggcggctccg gtggctccta atctagaact atagtgagtc
     1141 gtattacgta gatccagaca tgataagata cattgatgag tttggacaaa ccacaactag
     1201 aatgcagtga aaaaaatgct ttatttgtga aatttgtgat gctattgctt tatttgtaac
     1261 cattataagc tgcaataaac aagttaacaa caacaattgc attcatttta tgtttcaggt
     1321 tcagggggag gtgtgggagg ttttttaatt cctcaagaga cctctagacc agccaggaca
     1381 gaaatgcctc gacttcgctg ctacccaagg ttgccgggtg acgcacaccg tggaaacgga
     1441 tgaaggcacg aacccagtgg acataagcct gttcggttcg taagctgtaa tgcaagtagc
     1501 gtatgcgctc acgcaactgg tccagaacct tgaccgaacg cagcggtggt aacggcgcag
     1561 tggcggtttt catggcttgt tatgactgtt tttttggggt acagtctatg cctcgggcat
     1621 ccaagcagca agcgcgttac gccgtgggtc gatgtttgat gttatggagc agcaacgatg
     1681 ttacgcagca gggcagtcgc cctaaaacaa agttaaacat tatgagggaa gcggtgatcg
     1741 ccgaagtatc gactcaacta tcagaggtag ttggcgtcat cgagcgccat ctcgaaccga
     1801 cgttgctggc cgtacatttg tacggctccg cagtggatgg cggcctgaag ccacacagtg
     1861 atattgattt gctggttacg gtgaccgtaa ggcttgatga aacaacgcgg cgagctttga
     1921 tcaacgacct tttggaaact tcggcttccc ctggagagag cgagattctc cgcgctgtag
     1981 aagtcaccat tgttgtgcac gacgacatca ttccgtggcg ttatccagct aagcgcgaac
     2041 tgcaatttgg agaatggcag cgcaatgaca ttcttgcagg tatcttcgag ccagccacga
     2101 tcgacattga tctggctatc ttgctgacaa aagcaagaga acatagcgtt gccttggtag
     2161 gtccagcggc ggaggaactc tttgatccgg ttcctgaaca ggatctattt gaggcgctaa
     2221 atgaaacctt aacgctatgg aactcgccgc ccgactgggc tggcgatgag cgaaatgtag
     2281 tgcttacgtt gtcccgcatt tggtacagcg cagtaaccgg caaaatcgcg ccgaaggatg
     2341 tcgctgccga ctgggcaatg gagcgcctgc cggcccagta tcagcccgtc atacttgaag
     2401 ctagacaggc ttatcttgga caagaagaag atcgcttggc ctcgcgcgca gatcagttgg
     2461 aagaatttgt ccactacgtg aaaggcgaga tcaccaaggt agtcggcaaa taaccctcga
     2521 gccacccatg accaaaatcc cttaacgtga gttacgcgtc gttccactga gcgtcagacc
     2581 ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct
     2641 tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa
     2701 ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact gttcttctag
     2761 tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc
     2821 tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg
     2881 actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca
     2941 cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat
     3001 gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg
     3061 tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc
     3121 ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc
     3181 ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc
     3241 cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg
     3301 cctttgagtg agctgatacc gctcgccgca gccgaacgac cgagcgcagc gagtcagtga
     3361 gcgaggaagc ggaagagcgc ccaatacgca aaccgcctct ccccgcgcgt tggccgattc
     3421 attaatgcag ctggcacgac aggtttcccg actggaaagc gggcagtgag cgcaacgcaa
     3481 ttaatacgcg taccgctagc caggaagagt ttgtagaaac gcaaaaaggc catccgtcag
     3541 gatggccttc tgcttagttt gatgcctggc agtttatggc gggcgtcctg cccgccaccc
     3601 tccgggccgt tgcttcacaa cgttcaaatc cgctcccggc gg
//
