LOCUS       Exported                4302 bp ds-DNA     circular SYN 08-MAR-2024
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4302)
  TITLE     Shinoda H, et al. Acid-tolerant monomeric GFP from Olindias formosa.
            Cell Chem. Biol. 2018;25:330?338.
REFERENCE   2  (bases 1 to 4302)
  AUTHORS   SnapGene License
  TITLE     Direct Submission
  JOURNAL   Exported Monday, Jul 22, 2024 from SnapGene 5.0.8
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..4302
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     terminator      268..295
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      387..473
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     537..553
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(624..747)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     misc_feature    749..1335
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(916..933)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     1143..1162
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             1336..1338
                     /codon_start=1
                     /label=MET
                     /translation="M"
     CDS             1339..1380
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     primer_bind     1339..1358
                     /label=V5-FWD
     primer_bind     complement(1362..1380)
                     /label=V5-REV
     CDS             1381..1392
                     /codon_start=1
                     /label=MCS
                     /translation="GSTG"
     CDS             1393..1451
                     /codon_start=1
                     /label=GGGGS(x3)
                     /translation="EFDGGGGSGGGGSGGGGSP"
     CDS             1453..2160
                     /codon_start=1
                     /label=(n2)oxStayGold(c4)_v2
                     /translation="MVSKGEELFTGVTPFKFQLKGTINGKSFTVEGEGEGNSHEGSHKG
                     KYVCTSGKLPMSWAALGTSFGYGMKYYTKYPSGLKNWFHEVMPEGFTYDRHIQYKGDGS
                     IHAKHQHFMKNGTYHNIVEFTGQDFKENSPVLTGDMNVSLPNEVQHIPRDDGVECPVTL
                     LYPLLSDKSKCVEAYQNTIIKPLHNQPAPDVPYHWIRKQYTQSKDDTEERDHIIQSETL
                     EAHLPWHEPSASAV"
     misc_feature    1462..1488
                     /product="9 amino terminal AA from EGFP N2"
                     /label=n2
                     /note="EGFP aa 4-12"
     misc_feature    1489..2130
                     /label=StayGold (aa 4-X)
                     /note="missing amino terminal MAS of MAST sequence"
     misc_feature    1998..2001
                     /label=Cys>Ile
                     /note="cys->Ile for stability"
     misc_feature    2101..2103
                     /label=Cys>Ile
                     /note="Cys->Ile for stability"
     misc_feature    2131..2160
                     /label=C-term of dfGFP
                     /note="C-terminal of acid stable GFP (aa 223?233)"
     misc_feature    2133
                     /label=a>c
                     /note="to destroy NcoI site"
     polyA_signal    2208..2342
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     protein_bind    complement(2411..2506)
                     /gene="mutant version of attL"
                     /label=attL3
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        complement(2544..2562)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2567..2583)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             2696..3505
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     rep_origin      3652..4240
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga
       61 taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga
      121 gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca
      181 cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaata cgcgtaccgc
      241 tagccaggaa gagtttgtag aaacgcaaaa aggccatccg tcaggatggc cttctgctta
      301 gtttgatgcc tggcagttta tggcgggcgt cctgcccgcc accctccggg ccgttgcttc
      361 acaacgttca aatccgctcc cggcggattt gtcctactca ggagagcgtt caccgacaaa
      421 caacagataa aacgaaaggc ccagtcttcc gactgagcct ttcgttttat ttgatgcctg
      481 gcagttccct actctcgcgt taacgctagc atggatgttt tcccagtcac gacgttgtaa
      541 aacgacggcc agtcttaagc tcgggccctg cagctctaga gctcgaattc tacaggtcac
      601 taataccatc taagtagttg attcatagtg actgcatatg ttgtgtttta cagtattatg
      661 tagtctgttt tttatgcaaa atctaattta atatattgat atttatatca ttttacgttt
      721 ctcgttcaac tttcttgtac aaagtggccc cctctccctc ccccccccct aacgttactg
      781 gccgaagccg cttggaataa ggccggtgtg cgtttgtcta tatgttattt tccaccatat
      841 tgccgtcttt tggcaatgtg agggcccgga aacctggccc tgtcttcttg acgagcattc
      901 ctaggggtct ttcccctctc gccaaaggaa tgcaaggtct gttgaatgtc gtgaaggaag
      961 cagttcctct ggaagcttct tgaagacaaa caacgtctgt agcgaccctt tgcaggcagc
     1021 ggaacccccc acctggcgac aggtgcctct gcggccaaaa gccacgtgta taagatacac
     1081 ctgcaaaggc ggcacaaccc cagtgccacg ttgtgagttg gatagttgtg gaaagagtca
     1141 aatggctctc ctcaagcgta ttcaacaagg ggctgaagga tgcccagaag gtaccccatt
     1201 gtatgggatc tgatctgggg cctcggtaca catgctttac atgtgtttag tcgaggttaa
     1261 aaaaacgtct aggccccccg aaccacgggg acgtggtttt cctttgaaaa acacgatgat
     1321 aatatggcca caaccatggg taagcctatc cctaaccctc tcctcggtct cgattctacg
     1381 ggatccaccg gtgaattcga tggtggaggc ggttcaggcg gaggtggctc tggcggtggc
     1441 ggatcgccat caatggtgag caagggcgag gagctgttta caggcgtgac ccccttcaag
     1501 ttccagctga agggcaccat caacggcaag agcttcaccg tggaaggcga gggcgagggc
     1561 aatagccacg agggcagcca caaaggcaag tacgtgtgca ccagcggcaa actgccaatg
     1621 tcttgggccg ccctgggaac tagcttcggc tatggcatga agtactacac caagtacccc
     1681 agcggcctga agaactggtt ccacgaggtg atgcccgagg gcttcaccta cgacagacac
     1741 atccagtaca agggcgacgg cagcatccac gccaagcacc agcacttcat gaagaacggc
     1801 acctaccaca acatcgtgga gttcaccggc caggacttca aggagaacag ccccgtgctg
     1861 accggcgaca tgaacgtgag cctgcccaac gaggtgcagc acatccccag agatgacggc
     1921 gtggagtgcc cagtgaccct gctgtaccct ctgctgagcg acaagagcaa gtgcgtggag
     1981 gcctaccaga acaccatcat caagcccctg cacaatcagc cagcccccga tgtgccatac
     2041 cactggatca gaaagcagta cacccagagc aaggacgaca ccgaggagag agaccacatc
     2101 atccagagcg agaccctgga ggcccacctg ccctggcacg agccttctgc ttctgccgtg
     2161 tgatatctat aatgaggcgc gccgtcgaca cgcgttaatg atacgtagat ccagacatga
     2221 taagatacat tgatgagttt ggacaaacca caactagaat gcagtgaaaa aaatgcttta
     2281 tttgtgaaat ttgtgatgct attgctttat ttgtaaccat tataagctgc aataaacaag
     2341 ttaacaacaa caattgtcta gacattcatt ttatgtttca ggttcagggg gaggtgtggg
     2401 aggttttttc caactttatt atacaaagtt ggcattataa aaaagcattg cttatcaatt
     2461 tgttgcaacg aacaggtcac tatcagtcaa aataaaatca ttatttggag ctccatggta
     2521 gcgttaacgc ggccgcgata tcccctatag tgagtcgtat tacatggtca tagctgtttc
     2581 ctggcagctc tggcccgtgt ctcaaaatct ctgatgttac attgcacaag ataaaaatat
     2641 atcatcatga acaataaaac tgtctgctta cataaacagt aatacaaggg gtgttatgag
     2701 ccatattcaa cgggaaacgt cgaggccgcg attaaattcc aacatggatg ctgatttata
     2761 tgggtataaa tgggctcgcg ataatgtcgg gcaatcaggt gcgacaatct atcgcttgta
     2821 tgggaagccc gatgcgccag agttgtttct gaaacatggc aaaggtagcg ttgccaatga
     2881 tgttacagat gagatggtca gactaaactg gctgacggaa tttatgcctc ttccgaccat
     2941 caagcatttt atccgtactc ctgatgatgc atggttactc accactgcga tccccggaaa
     3001 aacagcattc caggtattag aagaatatcc tgattcaggt gaaaatattg ttgatgcgct
     3061 ggcagtgttc ctgcgccggt tgcattcgat tcctgtttgt aattgtcctt ttaacagcga
     3121 tcgcgtattt cgtctcgctc aggcgcaatc acgaatgaat aacggtttgg ttgatgcgag
     3181 tgattttgat gacgagcgta atggctggcc tgttgaacaa gtctggaaag aaatgcataa
     3241 acttttgcca ttctcaccgg attcagtcgt cactcatggt gatttctcac ttgataacct
     3301 tatttttgac gaggggaaat taataggttg tattgatgtt ggacgagtcg gaatcgcaga
     3361 ccgataccag gatcttgcca tcctatggaa ctgcctcggt gagttttctc cttcattaca
     3421 gaaacggctt tttcaaaaat atggtattga taatcctgat atgaataaat tgcagtttca
     3481 tttgatgctc gatgagtttt tctaatcaga attggttaat tggttgtaac actggcagag
     3541 cattacgctg acttgacggg acggcgcaag ctcatgacca aaatccctta acgtgagtta
     3601 cgcgtcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc
     3661 tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt
     3721 ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc
     3781 gcagatacca aatactgtcc ttctagtgta gccgtagtta ggccaccact tcaagaactc
     3841 tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg
     3901 cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg
     3961 gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga
     4021 actgagatac ctacagcgtg agcattgaga aagcgccacg cttcccgaag ggagaaaggc
     4081 ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg
     4141 gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg
     4201 atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt
     4261 tttacggttc ctggcctttt gctggccttt tgctcacatg tt
//
