LOCUS       Exported                3493 bp ds-DNA     circular SYN 12-AUG-2022
DEFINITION  .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3493)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Wednesday, Apr 5, 2023 from SnapGene 4.3.11
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..3493
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     terminator      1..87
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
                     /note="color: #ffffff"
     primer_bind     21..40
                     /label=A423 rrnb T1 ter -f
                     /note="color: black; sequence: cgttcaccgacaaacaacag; added:
                     2022-07-25"
     primer_bind     127..146
                     /label=C133 pME 5' con -f
                     /note="color: black; sequence: ggatgttttcccagtcacga; added:
                     2022-07-25"
     primer_bind     136..158
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
                     /note="color: black; sequence: CCCAGTCACGACGTTGTAAAACG"
     primer_bind     150..167
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
                     /note="color: black; sequence: TGTAAAACGACGGCCAGT"
     primer_bind     151..167
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
                     /note="color: #a020f0; direction: RIGHT"
     primer_bind     182..216
                     /label=P2A-BsaI-AGTC-ORF-1-f
                     /note="color: black; sequence: 
                     atcggtctctAGTCgctacaaacttctctctgctg; added: 2022-08-02"
     primer_bind     182..216
                     /label=P2A-BsaI-AGTC-ORF-2-f
                     /note="color: black; sequence: 
                     atcggtctctAGTCcgctacaaacttctctctgctg; added: 2022-08-02"
     primer_bind     182..216
                     /label=P2A-BsaI-AGTC-ORF-3-f
                     /note="color: black; sequence: 
                     atcggtctctAGTCccgctacaaacttctctctgctg; added: 2022-08-02"
     CDS             196..252
                     /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."
                     /note="color: #cc99b2
                     Cleavage site after base 250"
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             253..972
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /note="This feature has 3 segments:
                        1: 253 .. 255 / #05fd14
                        2: 256 .. 258 / #05fd14 / 1a
                        3: 259 .. 972 / #05fd14"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGGTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     253..273
                     /label=A605 GFP -f
                     /note="color: black; sequence: atggtgagcaagggcgaggag; 
                     added: 2022-08-10"
     primer_bind     253..272
                     /label=kozak-EGFP-BsaI-AGTC-f
                     /note="color: black; sequence: 
                     atcggtctctAGTCgccaccatggtgagcaagggcgagga; added: 
                     2022-08-02"
     primer_bind     complement(298..319)
                     /label=A228 GFP-N
                     /note="color: black; sequence: cgtcgccgtccagctcgaccag"
     primer_bind     complement(731..751)
                     /label=A197 C19 GFP confirm -r
                     /note="color: black; sequence: tgaagttcaccttgatgccgt"
     primer_bind     893..912
                     /label=A875 GFP confirm -f
                     /note="color: black; sequence: agaagcgcgatcacatggtc; added:
                     2022-08-10"
     promoter        complement(980..997)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
                     /note="color: #ffffff; direction: LEFT"
     polyA_signal    1002..1136
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
                     /note="color: #a6acb3"
     primer_bind     complement(1180..1213)
                     /label=SV40-pA-BsaI-CTCA-r
                     /note="color: black; sequence: 
                     gatggtctctTGAGgaattaaaaaacctcccacacct; added: 2022-08-02"
     CDS             1573..2364
                     /codon_start=1
                     /gene="aadA"
                     /product="aminoglycoside adenylyltransferase (Murphy, 
                     1985)"
                     /label=SmR
                     /note="confers resistance to spectinomycin and 
                     streptomycin"
                     /note="color: #ccffcc"
                     /translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
                     SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
                     RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
                     EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
                     VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK"
     rep_origin      2457..3045
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
                     /note="color: #ffff00"
     primer_bind     2946..2965
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
                     /note="color: black; sequence: GGGAAACGCCTGGTATCTTT"
     primer_bind     3199..3216
                     /label=A757 L4440
                     /note="L4440 vector, forward primer"
                     /note="color: black; sequence: AGCGAGTCAGTGAGCGAG"
     terminator      3375..3402
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
                     /note="color: #ffffff"
ORIGIN
        1 atttgtccta ctcaggagag cgttcaccga caaacaacag ataaaacgaa aggcccagtc
       61 ttccgactga gcctttcgtt ttatttgatg cctggcagtt ccctactctc gcgttaacgc
      121 tagcatggat gttttcccag tcacgacgtt gtaaaacgac ggccagtctt aagctcgggc
      181 ccggtctcta gtcccgctac aaacttctct ctgctgaagc aggctggaga cgtggaggag
      241 aaccctggac ctatggtgag caagggcgag gagctgttca ccggggtggt gcccatcctg
      301 gtcgagctgg acggcgacgt aaacggccac aagttcagtg tgtccggcga gggcgagggc
      361 gatgccacct acggcaagct gaccctgaag ttcatctgca ccaccggcaa gctgcccgtg
      421 ccctggccca ccctcgtgac caccctgacc tacggcgtgc agtgcttcag ccgctacccc
      481 gaccacatga agcagcacga cttcttcaag tccgccatgc ccgaaggcta cgtccaggag
      541 cgcaccatct tcttcaagga cgacggcaac tacaagaccc gcgccgaggt gaagttcgag
      601 ggcggcaccc tggtgaaccg catcgagctg aagggcatcg acttcaagga ggacggcaac
      661 atcctggggc ataagctgga gtacaactac aacagccaca acgtctatat catggccgac
      721 aagcagaaga acggcatcaa ggtgaacttc aagatccgcc acaacatcga ggacggcagc
      781 gtgcagctcg ccgaccacta ccagcagaac acccccatcg gcgacggccc cgtgctgctg
      841 cccgacaacc actacctgag cacccagtcc gccctgagca aagaccccaa cgagaagcgc
      901 gatcacatgg tcctgctgga gttcgtgacc gccgccggga tcactctcgg catggacgag
      961 ctgtacaagt aagctagaac tatagtgagt cgtattacgt agatccagac atgataagat
     1021 acattgatga gtttggacaa accacaacta gaatgcagtg aaaaaaatgc tttatttgtg
     1081 aaatttgtga tgctattgct ttatttgtaa ccattataag ctgcaataaa caagttaaca
     1141 acaacaattg cattcatttt atgtttcagg ttcaggggga ggtgtgggag gttttttaat
     1201 tcctcaagag acctctagac cagccaggac agaaatgcct cgacttcgct gctacccaag
     1261 gttgccgggt gacgcacacc gtggaaacgg atgaaggcac gaacccagtg gacataagcc
     1321 tgttcggttc gtaagctgta atgcaagtag cgtatgcgct cacgcaactg gtccagaacc
     1381 ttgaccgaac gcagcggtgg taacggcgca gtggcggttt tcatggcttg ttatgactgt
     1441 ttttttgggg tacagtctat gcctcgggca tccaagcagc aagcgcgtta cgccgtgggt
     1501 cgatgtttga tgttatggag cagcaacgat gttacgcagc agggcagtcg ccctaaaaca
     1561 aagttaaaca ttatgaggga agcggtgatc gccgaagtat cgactcaact atcagaggta
     1621 gttggcgtca tcgagcgcca tctcgaaccg acgttgctgg ccgtacattt gtacggctcc
     1681 gcagtggatg gcggcctgaa gccacacagt gatattgatt tgctggttac ggtgaccgta
     1741 aggcttgatg aaacaacgcg gcgagctttg atcaacgacc ttttggaaac ttcggcttcc
     1801 cctggagaga gcgagattct ccgcgctgta gaagtcacca ttgttgtgca cgacgacatc
     1861 attccgtggc gttatccagc taagcgcgaa ctgcaatttg gagaatggca gcgcaatgac
     1921 attcttgcag gtatcttcga gccagccacg atcgacattg atctggctat cttgctgaca
     1981 aaagcaagag aacatagcgt tgccttggta ggtccagcgg cggaggaact ctttgatccg
     2041 gttcctgaac aggatctatt tgaggcgcta aatgaaacct taacgctatg gaactcgccg
     2101 cccgactggg ctggcgatga gcgaaatgta gtgcttacgt tgtcccgcat ttggtacagc
     2161 gcagtaaccg gcaaaatcgc gccgaaggat gtcgctgccg actgggcaat ggagcgcctg
     2221 ccggcccagt atcagcccgt catacttgaa gctagacagg cttatcttgg acaagaagaa
     2281 gatcgcttgg cctcgcgcgc agatcagttg gaagaatttg tccactacgt gaaaggcgag
     2341 atcaccaagg tagtcggcaa ataaccctcg agccacccat gaccaaaatc ccttaacgtg
     2401 agttacgcgt cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga
     2461 gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg
     2521 gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc
     2581 agagcgcaga taccaaatac tgtccttcta gtgtagccgt agttaggcca ccacttcaag
     2641 aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc
     2701 agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg
     2761 cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac
     2821 accgaactga gatacctaca gcgtgagcat tgagaaagcg ccacgcttcc cgaagggaga
     2881 aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt
     2941 ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag
     3001 cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg
     3061 gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta
     3121 tcccctgatt ctgtggataa ccgtattacc gcctttgagt gagctgatac cgctcgccgc
     3181 agccgaacga ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg cccaatacgc
     3241 aaaccgcctc tccccgcgcg ttggccgatt cattaatgca gctggcacga caggtttccc
     3301 gactggaaag cgggcagtga gcgcaacgca attaatacgc gtaccgctag ccaggaagag
     3361 tttgtagaaa cgcaaaaagg ccatccgtca ggatggcctt ctgcttagtt tgatgcctgg
     3421 cagtttatgg cgggcgtcct gcccgccacc ctccgggccg ttgcttcaca acgttcaaat
     3481 ccgctcccgg cgg
//
