LOCUS       sequence-482081-        3431 bp    DNA     circular SYN 05-AUG-2025
DEFINITION  Middle entry vector for mini-Golden subcloning platform; mNeongreen.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3431)
  TITLE     Mini-Golden Subcloning Platform
REFERENCE   2  (bases 1 to 3431)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Aug 5, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"
COMMENT     Sequence Label: pMid-TCGC-mNeongreen
FEATURES             Location/Qualifiers
     source          1..3431
                     /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"
     regulatory      194..203
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             200..910
                     /codon_start=1
                     /product="bright monomeric yellow-green fluorescent protein
                     derived from LanYFP (Shaner et al., 2013)"
                     /label=mNeonGreen
                     /translation="MVSKGEEDNMASLPATHELHIFGSINGVDFDMVGQGTGNPNDGYE
                     ELNLKSTKGDLQFSPWILVPHIGYGFHQYLPYPDGMSPFQAAMVDGSGYQVHRTMQFED
                     GASLTVNYRYTYEGSHIKGEAQVKGTGFPADGPVMTNSLTAADWCRSKKTYPNDKTIIS
                     TFKWSYTTGNGKRYRSTARTTYTFAKPMAANYLKNQPMYVFRKTELKHSKTELNFKEWQ
                     KAFTDVMGMDELYK"
     regulatory      455..464
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     polyA_signal    940..1074
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(964..983)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1018..1037
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             1511..2302
                     /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      2395..2983
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2884..2903
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3137..3154
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(3313..3340)
                     /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 cgcgccacca tggtgagcaa gggcgaggag gataacatgg cctctctccc
      241 agcgacacat gagttacaca tctttggctc catcaacggt gtggactttg acatggtggg
      301 tcagggcacc ggcaatccaa atgatggtta tgaggagtta aacctgaagt ccaccaaggg
      361 tgacctccag ttctccccct ggattctggt ccctcatatc gggtatggct tccatcagta
      421 cctgccctac cctgacggga tgtcgccttt ccaggccgcc atggtagatg gctccggcta
      481 ccaagtccat cgcacaatgc agtttgaaga tggtgcctcc cttactgtta actaccgcta
      541 cacctacgag ggaagccaca tcaaaggaga ggcccaggtg aaggggactg gtttccctgc
      601 tgacggtcct gtgatgacca actcgctgac cgctgcggac tggtgcaggt cgaagaagac
      661 ttaccccaac gacaaaacca tcatcagtac ctttaagtgg agttacacca ctggaaatgg
      721 caagcgctac cggagcactg cgcggaccac ctacaccttt gccaagccaa tggcggctaa
      781 ctatctgaag aaccagccga tgtacgtgtt ccgtaagacg gagctcaagc actccaagac
      841 cgagctcaac ttcaaggagt ggcaaaaggc ctttaccgat gtgatgggca tggacgagct
      901 gtacaagtaa gctagaacta tagtgagtcg tattacgtag atccagacat gataagatac
      961 attgatgagt ttggacaaac cacaactaga atgcagtgaa aaaaatgctt tatttgtgaa
     1021 atttgtgatg ctattgcttt atttgtaacc attataagct gcaataaaca agttaacaac
     1081 aacaattgca ttcattttat gtttcaggtt cagggggagg tgtgggaggt tttttaattc
     1141 ctcaagagac ctctagacca gccaggacag aaatgcctcg acttcgctgc tacccaaggt
     1201 tgccgggtga cgcacaccgt ggaaacggat gaaggcacga acccagtgga cataagcctg
     1261 ttcggttcgt aagctgtaat gcaagtagcg tatgcgctca cgcaactggt ccagaacctt
     1321 gaccgaacgc agcggtggta acggcgcagt ggcggttttc atggcttgtt atgactgttt
     1381 ttttggggta cagtctatgc ctcgggcatc caagcagcaa gcgcgttacg ccgtgggtcg
     1441 atgtttgatg ttatggagca gcaacgatgt tacgcagcag ggcagtcgcc ctaaaacaaa
     1501 gttaaacatt atgagggaag cggtgatcgc cgaagtatcg actcaactat cagaggtagt
     1561 tggcgtcatc gagcgccatc tcgaaccgac gttgctggcc gtacatttgt acggctccgc
     1621 agtggatggc ggcctgaagc cacacagtga tattgatttg ctggttacgg tgaccgtaag
     1681 gcttgatgaa acaacgcggc gagctttgat caacgacctt ttggaaactt cggcttcccc
     1741 tggagagagc gagattctcc gcgctgtaga agtcaccatt gttgtgcacg acgacatcat
     1801 tccgtggcgt tatccagcta agcgcgaact gcaatttgga gaatggcagc gcaatgacat
     1861 tcttgcaggt atcttcgagc cagccacgat cgacattgat ctggctatct tgctgacaaa
     1921 agcaagagaa catagcgttg ccttggtagg tccagcggcg gaggaactct ttgatccggt
     1981 tcctgaacag gatctatttg aggcgctaaa tgaaacctta acgctatgga actcgccgcc
     2041 cgactgggct ggcgatgagc gaaatgtagt gcttacgttg tcccgcattt ggtacagcgc
     2101 agtaaccggc aaaatcgcgc cgaaggatgt cgctgccgac tgggcaatgg agcgcctgcc
     2161 ggcccagtat cagcccgtca tacttgaagc tagacaggct tatcttggac aagaagaaga
     2221 tcgcttggcc tcgcgcgcag atcagttgga agaatttgtc cactacgtga aaggcgagat
     2281 caccaaggta gtcggcaaat aaccctcgag ccacccatga ccaaaatccc ttaacgtgag
     2341 ttacgcgtcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga
     2401 tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt
     2461 ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag
     2521 agcgcagata ccaaatactg ttcttctagt gtagccgtag ttaggccacc acttcaagaa
     2581 ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag
     2641 tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca
     2701 gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac
     2761 cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa
     2821 ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc
     2881 agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg
     2941 tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc
     3001 ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc
     3061 ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg ctcgccgcag
     3121 ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc caatacgcaa
     3181 accgcctctc cccgcgcgtt ggccgattca ttaatgcagc tggcacgaca ggtttcccga
     3241 ctggaaagcg ggcagtgagc gcaacgcaat taatacgcgt accgctagcc aggaagagtt
     3301 tgtagaaacg caaaaaggcc atccgtcagg atggccttct gcttagtttg atgcctggca
     3361 gtttatggcg ggcgtcctgc ccgccaccct ccgggccgtt gcttcacaac gttcaaatcc
     3421 gctcccggcg g
//
