LOCUS       pKG2023_pGEEC509        4581 bp    DNA     circular SYN 25-APR-2023
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4581)
  AUTHORS   MIT
  TITLE     Direct Submission
  JOURNAL   Exported May 7, 2025 from SnapGene 8.0.3
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..4581
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1..42
                     /label=oMJ001
     primer_bind     1..42
                     /label=oMJ005
     primer_bind     3..42
                     /label=oMJ009
     primer_bind     3..42
                     /label=oMJ013
     primer_bind     9..39
                     /label=oMJ020
     primer_bind     complement(21..46)
                     /label=oSEQ033 Pentr_rev
     misc_feature    23..46
                     /label=pENTR2B.Seq.REV
     primer_bind     complement(35..56)
                     /label=oTA069 mIR-backbone_junction_rev
     primer_bind     complement(35..56)
                     /label=oSEQ044 pENTR_rev_seq
     primer_bind     complement(108..128)
                     /label=oSEQ005 pENTR_rev
                     /note="25nm	STD"
     CDS             122..931
                     /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
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPGDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(147..168)
                     /label=oSEQ023 attl2_rev
     primer_bind     complement(155..178)
                     /label=Kan_rev_SEQ
     primer_bind     complement(545..574)
                     /label=oTA099 GG_entry_upstream_fwd
     primer_bind     545..574
                     /label=oTA102 GG_entry_downstream_rev
     misc_feature    1005..1624
                     /label=pBR322_Origin
     rep_origin      1021..1609
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1416..1431
                     /label=oTA073 Xho_addition_rev
     primer_bind     complement(1486..1501)
                     /label=oTA072 Xho_addition_for
     primer_bind     1510..1529
                     /label=oSEQ067 pBR322-ori_fwd
     primer_bind     1761..1774
                     /label=oSEQ011 pEntr_for
                     /note="25nm	STD"
     terminator      1774..1860
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     1923..1942
                     /label=oSEQ011 pEntr_for
                     /note="25nm	STD"
     terminator      1952..1979
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     primer_bind     1981..2000
                     /label=pENTR_fwd_seq
     primer_bind     1981..2000
                     /label=oSEQ043 pENTR_fwd_seq
     primer_bind     complement(2002..2046)
                     /label=oMJ002
     primer_bind     complement(2002..2042)
                     /label=oMJ010
     primer_bind     complement(2002..2038)
                     /label=oMJ006
     primer_bind     complement(2002..2038)
                     /label=oMJ014
     primer_bind     complement(2010..2038)
                     /label=oMJ019
     primer_bind     2043..2064
                     /label=pPV1-UbC_fwd
     promoter        2047..3258
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     complement(2179..2198)
                     /label=oSEQ007
     primer_bind     complement(2179..2198)
                     /label=oSEQ007 UBC_rev
     primer_bind     2512..2531
                     /label=UbC-mut_fwd
     primer_bind     complement(2512..2531)
                     /label=UbC-mut_rev
     primer_bind     2870..2879
                     /label=oSEQ023 attl2_rev
     primer_bind     complement(2930..2946)
                     /label=oEP125
     primer_bind     2930..2946
                     /label=oEP126
     primer_bind     3096..3115
                     /label=oSEQ068 UbC_fwd
     primer_bind     complement(3221..3258)
                     /label=pPV1-UbC_rev
     primer_bind     complement(3236..3262)
                     /label=oEP127
     regulatory      3263..3272
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     3266..3287
                     /label=GG_mRuby2_fwd
     primer_bind     3268..3287
                     /label=mRuby2_fwd
                     /note="Fwd primer for mRuby2"
     CDS             3269..3979
                     /codon_start=1
                     /product="monomeric red fluorescent protein derived from 
                     mRuby, with improved photophysical properties (Lam et al., 
                     2012)"
                     /label=mRuby2
                     /translation="MVSKGEELIKENMRMKVVMEGSVNGHQFKCTGEGEGNPYMGTQTM
                     RIKVIEGGPLPFAFDILATSFMYGSRTFIKYPKGIPDFFKQSFPEGFTWERVTRYEDGG
                     VVTVMQDTSLEDGCLVYHVQVRGVNFPSNGPVMQKKTKGWEPNTEMMYPADGGLRGYTH
                     MALKVDGGGHLSCSFVTTYRSKKTVGNIKMPGIHAVDHRLERLEESDNEMFVVQREHAV
                     AKFAGLGGGMDELYK"
     primer_bind     complement(3295..3316)
                     /label=oEP076
     primer_bind     3317..3336
                     /label=oEP073
     primer_bind     complement(3964..3986)
                     /label=gemRuby2_rev
     primer_bind     complement(3964..3983)
                     /label=mRuby2_rev
                     /note="Rev primer for mRuby2"
     primer_bind     complement(3964..3983)
                     /label=GG_mRuby2_rev
     primer_bind     3982..4020
                     /label=geWPRE_fwd
     primer_bind     3987..4016
                     /label=GG_WPRE_fwd
     misc_feature    3993..4581
                     /label=WPRE
                     /note="woodchuck hepatitis virus 
                     posttranscriptionalregulatory element"
     primer_bind     complement(4159..4178)
                     /label=oSEQ034 WPRE_rev
     primer_bind     4416..4436
                     /label=oSEQ048 phage-dest-fwd-2
     primer_bind     complement(join(4564..4581,1..4))
                     /label=geWPRE_v2_rev
     primer_bind     complement(4564..4581)
                     /label=geWPRE_rev
     primer_bind     complement(4572..4581)
                     /label=GG_WPRE_rev
ORIGIN
        1 cgctcgctga ctgcaggtgc ccgtgtctca aaatctctga tgttacattg cacaagataa
       61 aaatatatca tcatgaacaa taaaactgtc tgcttacata aacagtaata caaggggtgt
      121 tatgagccat attcaacggg aaacgtcgag gccgcgatta aattccaaca tggatgctga
      181 tttatatggg tataaatggg ctcgcgataa tgtcgggcaa tcaggtgcga caatctatcg
      241 cttgtatggg aagcccgatg cgccagagtt gtttctgaaa catggcaaag gtagcgttgc
      301 caatgatgtt acagatgaga tggtcagact aaactggctg acggaattta tgcctcttcc
      361 gaccatcaag cattttatcc gtactcctgg tgatgcatgg ttactcacca ctgcgatccc
      421 cggaaaaaca gcattccagg tattagaaga atatcctgat tcaggtgaaa atattgttga
      481 tgcgctggca gtgttcctgc gccggttgca ttcgattcct gtttgtaatt gtccttttaa
      541 cagcgatcgc gtatttcgtc tagctcaggc gcaatcacga atgaataacg gtttggttga
      601 tgcgagtgat tttgatgacg agcgtaatgg ctggcctgtt gaacaagtct ggaaagaaat
      661 gcataaactt ttgccattct caccggattc agtcgtcact catggtgatt tctcacttga
      721 taaccttatt tttgacgagg ggaaattaat aggttgtatt gatgttggac gagtcggaat
      781 cgcagaccga taccaggatc ttgccatcct atggaactgc ctcggtgagt tttctccttc
      841 attacagaaa cggctttttc aaaaatatgg tattgataat cctgatatga ataaattgca
      901 gtttcatttg atgctcgatg agtttttcta atcagaattg gttaattggt tgtaacatta
      961 ttcagattgg gccccgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc
     1021 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
     1081 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
     1141 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
     1201 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
     1261 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
     1321 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
     1381 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
     1441 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
     1501 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
     1561 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
     1621 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
     1681 gttatcccct gattctgtgg ataaccgtat taccgctagc atggatctcg gggacgtcta
     1741 actactaagc gagagtaggg aactgccagg catcaaataa aacgaaaggc tcagtcggaa
     1801 gactgggcct ttcgttttat ctgttgtttg tcggtgaacg ctctcctgag taggacaaat
     1861 ccgccgggag cggatttgaa cgttgtgaag caacggcccg gagggtggcg ggcaggacgc
     1921 ccgccataaa ctgccaggca tcaaactaag cagaaggcca tcctgacgga tggccttttt
     1981 gcgtttctac aaactcttcc tgttagttag ttacttaagc tcgggcccac ctgcagtcta
     2041 cttactggcc tccgcgccgg gttttggcgc ctcccgcggg cgcccccctc ctcacggcga
     2101 gcgctgccac gtcagacgaa gggcgcagcg agcgtcctga tccttccgcc cggacgctca
     2161 ggacagcggc ccgctgctca taagactcgg ccttagaacc ccagtatcag cagaaggaca
     2221 ttttaggacg ggacttgggt gactctaggg cactggtttt ctttccagag agcggaacag
     2281 gcgaggaaaa gtagtccctt ctcggcgatt ctgcggaggg atctccgtgg ggcggtgaac
     2341 gccgatgatt atataaggac gcgccgggtg tggcacagct agttccgtcg cagccgggat
     2401 ttgggtcgcg gttcttgttt gtggatcgct gtgatcgtca cttggtgagt agcgggctgc
     2461 tgggctggcc ggggctttcg tggccgccgg gccgctcggt gggacggaag cgtgtggaga
     2521 gatcgccaag ggctgtagtc tgggtccgcg agcaaggttg ccctgaactg ggggttgggg
     2581 ggagcgcagc aaaatggcgg ctgttcccga gtcttgaatg gaagacgctt gtgaggcggg
     2641 ctgtgaggtc gttgaaacaa ggtggggggc atggtgggcg gcaagaaccc aaggtcttga
     2701 ggccttcgct aatgcgggaa agctcttatt cgggtgagat gggctggggc accatctggg
     2761 gaccctgacg tgaagtttgt cactgactgg agaactcggt ttgtcgtctg ttgcgggggc
     2821 ggcagttatg gcggtgccgt tgggcagtgc acccgtacct ttgggagcgc gcgccctcgt
     2881 cgtgtcgtga cgtcacccgt tctgttggct tataatgcag ggtggggcca cctgtcggta
     2941 ggtgtgcggt aggcttttct ccgtcgcagg acgcagggtt cgggcctagg gtaggctctc
     3001 ctgaatcgac aggcgccgga cctctggtga ggggagggat aagtgaggcg tcagtttctt
     3061 tggtcggttt tatgtaccta tcttcttaag tagctgaagc tccggttttg aactatgcgc
     3121 tcggggttgg cgagtgtgtt ttgtgaagtt ttttaggcac cttttgaaat gtaatcattt
     3181 gggtcaatat gtaattttca gtgttagact agtaaattgt ccgctaaatt ctggccgttt
     3241 ttggcttttt tgttagacaa tggccaccat ggtgtctaag ggcgaagagc tgatcaagga
     3301 aaatatgcgt atgaaggtgg tcatggaagg ttcggtcaac ggccaccaat tcaaatgcac
     3361 aggtgaagga gaaggcaatc cgtacatggg aactcaaacc atgaggatca aagtcatcga
     3421 gggaggaccc ctgccatttg cctttgacat tcttgccacg tcgttcatgt atggcagccg
     3481 tacttttatc aagtacccga aaggcattcc tgatttcttt aaacagtcct ttcctgaggg
     3541 ttttacttgg gaaagagtta cgagatacga agatggtgga gtcgtcaccg tcatgcagga
     3601 caccagcctt gaggatggct gtctcgttta ccacgtccaa gtcagagggg taaactttcc
     3661 ctccaatggt cccgtgatgc agaagaagac caagggttgg gagcctaata cagagatgat
     3721 gtatccagca gatggtggtc tgaggggata cactcatatg gcactgaaag ttgatggtgg
     3781 tggccatctg tcttgctctt tcgtaacaac ttacaggtca aaaaagaccg tcgggaacat
     3841 caagatgccc ggtatccatg ccgttgatca ccgcctggaa aggttagagg aaagtgacaa
     3901 tgaaatgttc gtagtacaac gcgaacacgc agttgccaag ttcgccgggc ttggtggtgg
     3961 gatggacgag ctgtacaagt aacaacgccg ataatcaacc tctggattac aaaatttgtg
     4021 aaagattgac tggtattctt aactatgttg ctccttttac gctatgtgga tacgctgctt
     4081 taatgccttt gtatcatgct attgcttccc gtatggcttt cattttctcc tccttgtata
     4141 aatcctggtt gctgtctctt tatgaggagt tgtggcccgt tgtcaggcaa cgtggcgtgg
     4201 tgtgcactgt gtttgctgac gcaaccccca ctggttgggg cattgccacc acctgtcagc
     4261 tcctttccgg gactttcgct ttccccctcc ctattgccac ggcggaactc atcgccgcct
     4321 gccttgcccg ctgctggaca ggggctcggc tgttgggcac tgacaattcc gtggtgttgt
     4381 cggggaaatc atcgtccttt ccttggctgc tcgcctgtgt tgccacctgg attctgcgcg
     4441 ggacgtcctt ctgctacgtc ccttcggccc tcaatccagc ggaccttcct tcccgcggcc
     4501 tgctgccggc tctgcggcct cttccgcgtc ttcgccttcg ccctcagacg agtcggatct
     4561 ccctttgggc cgcctccccg c
//
