LOCUS       pKG3776_pGEEC583        3491 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 3491)
  AUTHORS   MIT
  TITLE     Direct Submission
  JOURNAL   Exported May 7, 2025 from SnapGene 8.0.3
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..3491
                     /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..115
                     /label=oKL206
     promoter        107..555
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     198..217
                     /label=hPGK-mut_fwd
     primer_bind     complement(198..217)
                     /label=hPGK-mut_rev
     primer_bind     complement(536..555)
                     /label=oEP194
     primer_bind     536..555
                     /label=oEP205 hPGK-rev
     primer_bind     536..554
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     primer_bind     complement(542..581)
                     /label=oEP196
     primer_bind     556..601
                     /label=oEP195
     misc_feature    556..581
                     /label=EFS 5' UTR
     primer_bind     559..578
                     /label=oEP209 EF1a-rev
     primer_bind     575..601
                     /label=oEP193
     regulatory      586..595
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     589..610
                     /label=GG_mRuby2_fwd
     primer_bind     591..610
                     /label=mRuby2_fwd
                     /note="Fwd primer for mRuby2"
     CDS             592..1302
                     /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     592..611
                     /label=oEP159
     primer_bind     complement(618..639)
                     /label=oEP076
     primer_bind     640..659
                     /label=oEP073
     primer_bind     662..681
                     /label=oSEQ062 mRuby2_fwd
     primer_bind     complement(662..681)
                     /label=oSEQ093 mRuby2_rev
     primer_bind     complement(1194..1213)
                     /label=oSEQ061 mRuby2_rev
     primer_bind     1228..1247
                     /label=OSEQ091 mRuby2_fwd_SEQ
     primer_bind     complement(1287..1314)
                     /label=GG_mRuby2_rev
     primer_bind     complement(1287..1309)
                     /label=gemRuby2_rev
     primer_bind     complement(1287..1306)
                     /label=mRuby2_rev
                     /note="Rev primer for mRuby2"
     primer_bind     1305..1329
                     /label=gebGH_fwd
     primer_bind     1311..1332
                     /label=GG_bGH_fwd
     primer_bind     1315..1332
                     /label=GG_WPRE-bGH_fwd
     polyA_signal    1317..1541
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(1401..1426)
                     /label=oEP181 IVT-bGH-rev
     misc_feature    1424..1426
                     /label=bGH TES
     primer_bind     complement(1523..1545)
                     /label=gebGH_rev
     primer_bind     complement(1524..1540)
                     /label=oSEQ078 bGH_rev
     primer_bind     complement(1527..1541)
                     /label=GG_bGH_rev
     primer_bind     1542..1583
                     /label=oMJ001
     primer_bind     1542..1583
                     /label=oMJ005
     primer_bind     1544..1583
                     /label=oMJ009
     primer_bind     1544..1583
                     /label=oMJ013
     primer_bind     1550..1580
                     /label=oMJ020
     primer_bind     complement(1562..1587)
                     /label=oSEQ033 Pentr_rev
     misc_feature    1564..1587
                     /label=pENTR2B.Seq.REV
     primer_bind     complement(1576..1597)
                     /label=oTA069 mIR-backbone_junction_rev
     primer_bind     complement(1576..1597)
                     /label=oSEQ044 pENTR_rev_seq
     primer_bind     complement(1649..1669)
                     /label=oSEQ005 pENTR_rev
                     /note="25nm	STD"
     CDS             1663..2472
                     /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(1688..1709)
                     /label=oSEQ023 attl2_rev
     primer_bind     complement(1696..1719)
                     /label=Kan_rev_SEQ
     primer_bind     complement(2086..2115)
                     /label=oTA099 GG_entry_upstream_fwd
     primer_bind     2086..2115
                     /label=oTA102 GG_entry_downstream_rev
     misc_feature    2546..3165
                     /label=pBR322_Origin
     rep_origin      2562..3150
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2594..2613
                     /label=oEP033
     primer_bind     complement(2594..2613)
                     /label=oEP035
     primer_bind     2914..2934
                     /label=oEP024
     primer_bind     complement(2914..2934)
                     /label=oEP025
     primer_bind     2957..2972
                     /label=oTA073 Xho_addition_rev
     primer_bind     complement(3027..3042)
                     /label=oTA072 Xho_addition_for
     primer_bind     3051..3070
                     /label=oSEQ067 pBR322-ori_fwd
     primer_bind     complement(3167..3186)
                     /label=oEP013
     primer_bind     3167..3186
                     /label=oEP014
     primer_bind     3302..3315
                     /label=oSEQ011 pEntr_for
                     /note="25nm	STD"
     terminator      3315..3401
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     3464..3483
                     /label=oSEQ011 pEntr_for
                     /note="25nm	STD"
ORIGIN
        1 cagaaggcca tcctgacgga tggccttttt gcgtttctac aaactcttcc tgttagttag
       61 ttacttaagc tcgggcccac ctgcagtcta cttactataa ggatccgggg ttggggttgc
      121 gccttttcca aggcagccct gggtttgcgc agggacgcgg ctgctctggg cgtggttccg
      181 ggaaacgcag cggcgccgac cctggatctc gcacattctt cacgtccgtt cgcagcgtca
      241 cccggatctt cgccgctacc cttgtgggcc ccccggcgac gcttcctgct ccgcccctaa
      301 gtcgggaagg ttccttgcgg ttcgcggcgt gccggacgtg acaaacggaa gccgcacgtc
      361 tcactagtac cctcgcagac ggacagcgcc agggagcaat ggcagcgcgc cgaccgcgat
      421 gggctgtggc caatagcggc tgctcagcag ggcgcgccga gagcagcggc cgggaagggg
      481 cggtgcggga ggcggggtgt ggggcggtag tgtgggccct gttcctgccc gcgcggtgtt
      541 ccgcattctg caagccaacg ggtttgccgc cagaacacag taatggccac catggtgtct
      601 aagggcgaag agctgatcaa ggaaaatatg cgtatgaagg tggtcatgga aggttcggtc
      661 aacggccacc aattcaaatg cacaggtgaa ggagaaggca atccgtacat gggaactcaa
      721 accatgagga tcaaagtcat cgagggagga cccctgccat ttgcctttga cattcttgcc
      781 acgtcgttca tgtatggcag ccgtactttt atcaagtacc cgaaaggcat tcctgatttc
      841 tttaaacagt cctttcctga gggttttact tgggaaagag ttacgagata cgaagatggt
      901 ggagtcgtca ccgtcatgca ggacaccagc cttgaggatg gctgtctcgt ttaccacgtc
      961 caagtcagag gggtaaactt tccctccaat ggtcccgtga tgcagaagaa gaccaagggt
     1021 tgggagccta atacagagat gatgtatcca gcagatggtg gtctgagggg atacactcat
     1081 atggcactga aagttgatgg tggtggccat ctgtcttgct ctttcgtaac aacttacagg
     1141 tcaaaaaaga ccgtcgggaa catcaagatg cccggtatcc atgccgttga tcaccgcctg
     1201 gaaaggttag aggaaagtga caatgaaatg ttcgtagtac aacgcgaaca cgcagttgcc
     1261 aagttcgccg ggcttggtgg tgggatggac gagctgtaca agtaacaacg ttgcacctgt
     1321 gccttctagt tgccagccat ctgttgtttg cccctccccc gtgccttcct tgaccctgga
     1381 aggtgccact cccactgtcc tttcctaata aaatgaggaa attgcatcgc attgtctgag
     1441 taggtgtcat tctattctgg ggggtggggt ggggcaggac agcaaggggg aggattggga
     1501 agacaatagc aggcatgctg gggatgcggt gggctctatg gcgctcgctg actgcaggtg
     1561 cccgtgtctc aaaatctctg atgttacatt gcacaagata aaaatatatc atcatgaaca
     1621 ataaaactgt ctgcttacat aaacagtaat acaaggggtg ttatgagcca tattcaacgg
     1681 gaaacgtcga ggccgcgatt aaattccaac atggatgctg atttatatgg gtataaatgg
     1741 gctcgcgata atgtcgggca atcaggtgcg acaatctatc gcttgtatgg gaagcccgat
     1801 gcgccagagt tgtttctgaa acatggcaaa ggtagcgttg ccaatgatgt tacagatgag
     1861 atggtcagac taaactggct gacggaattt atgcctcttc cgaccatcaa gcattttatc
     1921 cgtactcctg gtgatgcatg gttactcacc actgcgatcc ccggaaaaac agcattccag
     1981 gtattagaag aatatcctga ttcaggtgaa aatattgttg atgcgctggc agtgttcctg
     2041 cgccggttgc attcgattcc tgtttgtaat tgtcctttta acagcgatcg cgtatttcgt
     2101 ctagctcagg cgcaatcacg aatgaataac ggtttggttg atgcgagtga ttttgatgac
     2161 gagcgtaatg gctggcctgt tgaacaagtc tggaaagaaa tgcataaact tttgccattc
     2221 tcaccggatt cagtcgtcac tcatggtgat ttctcacttg ataaccttat ttttgacgag
     2281 gggaaattaa taggttgtat tgatgttgga cgagtcggaa tcgcagaccg ataccaggat
     2341 cttgccatcc tatggaactg cctcggtgag ttttctcctt cattacagaa acggcttttt
     2401 caaaaatatg gtattgataa tcctgatatg aataaattgc agtttcattt gatgctcgat
     2461 gagtttttct aatcagaatt ggttaattgg ttgtaacatt attcagattg ggccccgttc
     2521 cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg
     2581 cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg
     2641 gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca
     2701 aatactgttc ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg
     2761 cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg
     2821 tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga
     2881 acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac
     2941 ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat
     3001 ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc
     3061 tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga
     3121 tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc
     3181 ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg
     3241 gataaccgta ttaccgctag catggatctc ggggacgtct aactactaag cgagagtagg
     3301 gaactgccag gcatcaaata aaacgaaagg ctcagtcgga agactgggcc tttcgtttta
     3361 tctgttgttt gtcggtgaac gctctcctga gtaggacaaa tccgccggga gcggatttga
     3421 acgttgtgaa gcaacggccc ggagggtggc gggcaggacg cccgccataa actgccaggc
     3481 atcaaactaa g
//
