LOCUS       pKG3777_pGEEC584        3222 bp    DNA     circular SYN 23-JAN-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  recombinant plasmid
REFERENCE   1  (bases 1 to 3222)
  AUTHORS   MIT
  TITLE     Direct Submission
  JOURNAL   Exported May 7, 2025 from SnapGene 8.0.3
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..3222
                     /mol_type="other DNA"
                     /organism="recombinant plasmid"
     terminator      2..29
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     primer_bind     31..50
                     /label=pENTR_fwd_seq
     primer_bind     31..50
                     /label=oSEQ043 pENTR_fwd_seq
     primer_bind     complement(52..96)
                     /label=oMJ002
     primer_bind     complement(52..92)
                     /label=oMJ010
     primer_bind     complement(52..88)
                     /label=oMJ006
     primer_bind     complement(52..88)
                     /label=oMJ014
     primer_bind     complement(60..88)
                     /label=oMJ019
     primer_bind     93..113
                     /label=oKL204
     promoter        97..283
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     primer_bind     complement(139..156)
                     /label=oSEQ045 phage-dest-rev-2
     primer_bind     complement(263..312)
                     /label=oEP198
     primer_bind     264..283
                     /label=oEP207 EFS-rev
     primer_bind     284..332
                     /label=oEP193
     primer_bind     284..327
                     /label=oEP197
     misc_feature    284..312
                     /label=CMV 5' UTR
     regulatory      317..326
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     320..341
                     /label=GG_mRuby2_fwd
     primer_bind     322..341
                     /label=mRuby2_fwd
                     /note="Fwd primer for mRuby2"
     CDS             323..1033
                     /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     323..342
                     /label=oEP159
     primer_bind     complement(349..370)
                     /label=oEP076
     primer_bind     371..390
                     /label=oEP073
     primer_bind     393..412
                     /label=oSEQ062 mRuby2_fwd
     primer_bind     complement(393..412)
                     /label=oSEQ093 mRuby2_rev
     primer_bind     661..680
                     /label=oQT047 mRuby2_1_fwd
     primer_bind     784..803
                     /label=oQT049 mRuby2_2_fwd
     primer_bind     complement(822..841)
                     /label=oQT048 mRuby2_1_rev
     primer_bind     complement(924..943)
                     /label=oQT050 mRuby2_2_rev
     primer_bind     complement(925..944)
                     /label=oSEQ061 mRuby2_rev
     primer_bind     959..978
                     /label=OSEQ091 mRuby2_fwd_SEQ
     primer_bind     complement(1018..1045)
                     /label=GG_mRuby2_rev
     primer_bind     complement(1018..1040)
                     /label=gemRuby2_rev
     primer_bind     complement(1018..1037)
                     /label=mRuby2_rev
                     /note="Rev primer for mRuby2"
     primer_bind     1036..1060
                     /label=gebGH_fwd
     primer_bind     1042..1063
                     /label=GG_bGH_fwd
     primer_bind     1046..1063
                     /label=GG_WPRE-bGH_fwd
     polyA_signal    1048..1272
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(1132..1157)
                     /label=oEP181
                     /note="IVT reverse primer for bGH"
     misc_feature    1155..1157
                     /label=bGH TES
     primer_bind     complement(1254..1276)
                     /label=gebGH_rev
     primer_bind     complement(1255..1271)
                     /label=oSEQ078 bGH_rev
     primer_bind     complement(1258..1272)
                     /label=GG_bGH_rev
     primer_bind     1273..1314
                     /label=oMJ001
     primer_bind     1273..1314
                     /label=oMJ005
     primer_bind     1275..1314
                     /label=oMJ009
     primer_bind     1275..1314
                     /label=oMJ013
     primer_bind     1281..1311
                     /label=oMJ020
     primer_bind     complement(1293..1318)
                     /label=oSEQ033 Pentr_rev
     misc_feature    1295..1318
                     /label=pENTR2B.Seq.REV
     primer_bind     complement(1307..1328)
                     /label=oTA069 mIR-backbone_junction_rev
     primer_bind     complement(1307..1328)
                     /label=oSEQ044 pENTR_rev_seq
     primer_bind     complement(1380..1400)
                     /label=oSEQ005 pENTR_rev
                     /note="25nm	STD"
     CDS             1394..2203
                     /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(1419..1440)
                     /label=oSEQ023 attl2_rev
     primer_bind     complement(1427..1450)
                     /label=Kan_rev_SEQ
     primer_bind     complement(1817..1846)
                     /label=oTA099 GG_entry_upstream_fwd
     primer_bind     1817..1846
                     /label=oTA102 GG_entry_downstream_rev
     misc_feature    2277..2896
                     /label=pBR322_Origin
     rep_origin      2293..2881
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2325..2344
                     /label=oEP033
     primer_bind     complement(2325..2344)
                     /label=oEP035
     primer_bind     2645..2665
                     /label=oEP024
     primer_bind     complement(2645..2665)
                     /label=oEP025
     primer_bind     2688..2703
                     /label=oTA073 Xho_addition_rev
     primer_bind     complement(2758..2773)
                     /label=oTA072 Xho_addition_for
     primer_bind     2782..2801
                     /label=oSEQ067 pBR322-ori_fwd
     primer_bind     complement(2898..2917)
                     /label=oEP013
     primer_bind     2898..2917
                     /label=oEP014
     primer_bind     3033..3046
                     /label=oSEQ011 pEntr_for
                     /note="25nm	STD"
     terminator      3046..3132
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     3195..3214
                     /label=oSEQ011 pEntr_for
                     /note="25nm	STD"
ORIGIN
        1 cagaaggcca tcctgacgga tggccttttt gcgtttctac aaactcttcc tgttagttag
       61 ttacttaagc tcgggcccac ctgcagtcta cttactgggc agagcgcaca tcgcccacag
      121 tccccgagaa gttgggggga ggggtcggca attgaaccgg tgcctagaga aggtggcgcg
      181 gggtaaactg ggaaagtgat gtcgtgtact ggctccgcct ttttcccgag ggtgggggag
      241 aaccgtatat aagtgcagta gtcgccgtga acgttctttt tcgagatcgc ctggagacgc
      301 catccacgct gtaatggcca ccatggtgtc taagggcgaa gagctgatca aggaaaatat
      361 gcgtatgaag gtggtcatgg aaggttcggt caacggccac caattcaaat gcacaggtga
      421 aggagaaggc aatccgtaca tgggaactca aaccatgagg atcaaagtca tcgagggagg
      481 acccctgcca tttgcctttg acattcttgc cacgtcgttc atgtatggca gccgtacttt
      541 tatcaagtac ccgaaaggca ttcctgattt ctttaaacag tcctttcctg agggttttac
      601 ttgggaaaga gttacgagat acgaagatgg tggagtcgtc accgtcatgc aggacaccag
      661 ccttgaggat ggctgtctcg tttaccacgt ccaagtcaga ggggtaaact ttccctccaa
      721 tggtcccgtg atgcagaaga agaccaaggg ttgggagcct aatacagaga tgatgtatcc
      781 agcagatggt ggtctgaggg gatacactca tatggcactg aaagttgatg gtggtggcca
      841 tctgtcttgc tctttcgtaa caacttacag gtcaaaaaag accgtcggga acatcaagat
      901 gcccggtatc catgccgttg atcaccgcct ggaaaggtta gaggaaagtg acaatgaaat
      961 gttcgtagta caacgcgaac acgcagttgc caagttcgcc gggcttggtg gtgggatgga
     1021 cgagctgtac aagtaacaac gttgcacctg tgccttctag ttgccagcca tctgttgttt
     1081 gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc ctttcctaat
     1141 aaaatgagga aattgcatcg cattgtctga gtaggtgtca ttctattctg gggggtgggg
     1201 tggggcagga cagcaagggg gaggattggg aagacaatag caggcatgct ggggatgcgg
     1261 tgggctctat ggcgctcgct gactgcaggt gcccgtgtct caaaatctct gatgttacat
     1321 tgcacaagat aaaaatatat catcatgaac aataaaactg tctgcttaca taaacagtaa
     1381 tacaaggggt gttatgagcc atattcaacg ggaaacgtcg aggccgcgat taaattccaa
     1441 catggatgct gatttatatg ggtataaatg ggctcgcgat aatgtcgggc aatcaggtgc
     1501 gacaatctat cgcttgtatg ggaagcccga tgcgccagag ttgtttctga aacatggcaa
     1561 aggtagcgtt gccaatgatg ttacagatga gatggtcaga ctaaactggc tgacggaatt
     1621 tatgcctctt ccgaccatca agcattttat ccgtactcct ggtgatgcat ggttactcac
     1681 cactgcgatc cccggaaaaa cagcattcca ggtattagaa gaatatcctg attcaggtga
     1741 aaatattgtt gatgcgctgg cagtgttcct gcgccggttg cattcgattc ctgtttgtaa
     1801 ttgtcctttt aacagcgatc gcgtatttcg tctagctcag gcgcaatcac gaatgaataa
     1861 cggtttggtt gatgcgagtg attttgatga cgagcgtaat ggctggcctg ttgaacaagt
     1921 ctggaaagaa atgcataaac ttttgccatt ctcaccggat tcagtcgtca ctcatggtga
     1981 tttctcactt gataacctta tttttgacga ggggaaatta ataggttgta ttgatgttgg
     2041 acgagtcgga atcgcagacc gataccagga tcttgccatc ctatggaact gcctcggtga
     2101 gttttctcct tcattacaga aacggctttt tcaaaaatat ggtattgata atcctgatat
     2161 gaataaattg cagtttcatt tgatgctcga tgagtttttc taatcagaat tggttaattg
     2221 gttgtaacat tattcagatt gggccccgtt ccactgagcg tcagaccccg tagaaaagat
     2281 caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa
     2341 accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa
     2401 ggtaactggc ttcagcagag cgcagatacc aaatactgtt cttctagtgt agccgtagtt
     2461 aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt
     2521 accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata
     2581 gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt
     2641 ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac
     2701 gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga
     2761 gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg
     2821 ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa
     2881 aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat
     2941 gttctttcct gcgttatccc ctgattctgt ggataaccgt attaccgcta gcatggatct
     3001 cggggacgtc taactactaa gcgagagtag ggaactgcca ggcatcaaat aaaacgaaag
     3061 gctcagtcgg aagactgggc ctttcgtttt atctgttgtt tgtcggtgaa cgctctcctg
     3121 agtaggacaa atccgccggg agcggatttg aacgttgtga agcaacggcc cggagggtgg
     3181 cgggcaggac gcccgccata aactgccagg catcaaacta ag
//
