LOCUS       sequence-67102-e        8177 bp    DNA     circular SYN 14-APR-2025
DEFINITION  ecDHFR based destable domain function at low temperature for 
            regulating protein stability in Caenorhabditis elegans.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8177)
  AUTHORS   Cho U, Zimmerman SM, Chen LC, Owen E, Kim JV, Kim SK, Wandless TJ
  TITLE     Rapid and Tunable Control of Protein Stability in Caenorhabditis 
            elegans Using a Small Molecule.
  JOURNAL   PLoS One. 2013 Aug 22;8(8):e72393. doi: 
            10.1371/journal.pone.0072393.
  PUBMED    23991108
REFERENCE   2  (bases 1 to 8177)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "PLoS One.";
            date: "2013-08-22"; volume: "8(8):e72393. doi"; pages: " 
            10.1371/journal.pone.0072393"
COMMENT     Sequence Label: pBMN ltDHFR(DD)-YFP
FEATURES             Location/Qualifiers
     source          1..8177
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     LTR             1..594
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     misc_feature    657..1014
                     /label=MMLV Psi
                     /note="packaging signal of Moloney murine leukemia virus 
                     (MMLV)"
     CDS             1079..1495
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     1425..1447
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     1463..1479
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     misc_feature    1505..1878
                     /label=pol region
                     /note="Moloney murine leukemia virus (MMLV) pol region 
                     containing the splice acceptor site"
     primer_bind     1816..1832
                     /label=pBMN 5'
                     /note="MMLV sequence, for inserts in pBMN retroviral 
                     vector"
     CDS             1893..2369
                     /codon_start=1
                     /gene="folA"
                     /product="E. coli dihydrofolate reductase"
                     /label=ecDHFR
                     /translation="MISLIAALAVDHVIGMENAMPWSLPADLAWFKRNTLNKPVIMGRH
                     TWESIGRPLPGRKNIILSSQPSTDDRVTWVKSADEAIAACGDVPEIMVIGGGRVYEQFL
                     PKAQKLYLTHIDAEVGGDTHFPDYEPDDWGSVFSEFHDADAQNSHSYCFVILGRR"
     CDS             join(2376..2378,2379..3089)
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(2418..2439)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2679..2698)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3026..3047
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             3096..3122
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     misc_feature    join(3146..3720,3721..3723,3724..3732)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(3313..3330)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     3540..3559
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             3733..4416
                     /codon_start=1
                     /product="far-red fluorescent variant of Heteractis crispa 
                     chromoprotein"
                     /label=HcRed1
                     /note="mammalian codon-optimized"
                     /translation="MVSGLLKESMRIKMYMEGTVNGHYFKCEGEGDGNPFAGTQSMRIH
                     VTEGAPLPFAFDILAPCCEYGSRTFVHHTAEIPDFFKQSFPEGFTWERTTTYEDGGILT
                     AHQDTSLEGNCLIYKVKVHGTNFPADGPVMKNKSGGWEPSTEVVYPENGVLCGRNVMAL
                     KVGDRHLICHHYTSYRSKKAVRALTMPGFHFTDIRLQMLRKKKDEYFELYEASVARYSD
                     LPEKAN"
     CDS             4429..5115
                     /codon_start=1
                     /product="far-red fluorescent variant of Heteractis crispa 
                     chromoprotein"
                     /label=HcRed1
                     /note="mammalian codon-optimized"
                     /translation="MVSGLLKESMRIKMYMEGTVNGHYFKCEGEGDGNPFAGTQSMRIH
                     VTEGAPLPFAFDILAPCCEYGSRTFVHHTAEIPDFFKQSFPEGFTWERTTTYEDGGILT
                     AHQDTSLEGNCLIYKVKVHGTNFPADGPVMKNKSGGWEPSTEVVYPENGVLCGRNVMAL
                     KVGDRHLICHHYTSYRSKKAVRALTMPGFHFTDIRLQMLRKKKDEYFELYEASVARYSD
                     LPEKAN"
     LTR             5164..5757
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     primer_bind     complement(5888..5905)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6059..6647)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6139..6158)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6821..7612,7613..7681))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     7444..7463
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     primer_bind     7749..7767
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 aatgaaagac cccacctgta ggtttggcaa gctagcttaa gtaacgccat tttgcaaggc
       61 atggaaaaat acataactga gaatagaaaa gttcagatca aggtcaggaa cagatggaac
      121 agctgaatat gggccaaagc ggatatctgt ggtaagcagt tcctgccccg gctcagggcc
      181 aagaacagat ggaacagctg aatatgggcc aaacaggata tctgtggtaa gcagttcctg
      241 ccccggctca gggccaagaa cagatggtcc ccagatgcgg tccagccctc agcagtttct
      301 agagaaccat cagatgtttc cagggtgccc caaggacctg aaatgaccct gtgccttatt
      361 tgaactaacc aatcagttcg cttctcgctt ctgttcgcgc gcttctgctc cccgagctca
      421 ataaaagagc ccacaacccc tcactcgggg cgccagtcct ccgattgact gagtcgcccg
      481 ggtacccgtg tatccaataa accctcttgc agttgcatcc gacttgtggt ctcgctgttc
      541 cttgggaggg tctcctctga gtgattgact acccgtcagc gggggtcttt catttggggg
      601 ctcgtccggg atcgggagac ccctgcccag ggaccaccga cccaccaccg ggaggtaagc
      661 tggccagcaa cttatctgtg tctgtccgat tgtctagtgt ctatgactga ttttatgcgc
      721 ctgcgtcggt actagttagc taactagctc tgtatctggc ggacccgtgg tggaactgac
      781 gagttcggaa cacccggccg caaccctggg agacgtccca gggacttcgg gggccgtttt
      841 tgtggcccga cctgagtcca aaaatcccga tcgttttgga ctctttggtg cacccccctt
      901 agaggaggga tatgtggttc tggtaggaga cgagaaccta aaacagttcc cgcctccgtc
      961 tgaatttttg ctttcggttt gggaccgaag ccgcgccgcg cgtcttgtct gctgcagcat
     1021 cgttctgtgt tgtctctgtc tgactgtgtt tctgtatttg tctgaaaata tgggcccggg
     1081 ccagactgtt accactccct taagtttgac cttaggtcac tggaaagatg tcgagcggat
     1141 cgctcacaac cagtcggtag atgtcaagaa gagacgttgg gttaccttct gctctgcaga
     1201 atggccaacc tttaacgtcg gatggccgcg agacggcacc tttaaccgag acctcatcac
     1261 ccaggttaag atcaaggtct tttcacctgg cccgcatgga cacccagacc aggtccccta
     1321 catcgtgacc tgggaagcct tggcttttga cccccctccc tgggtcaagc cctttgtaca
     1381 ccctaagcct ccgcctcctc ttcctccatc cgccccgtct ctcccccttg aacctcctcg
     1441 ttcgaccccg cctcgatcct ccctttatcc agccctcact ccttctctag gcgcccccat
     1501 atggccatat gagatcttat atggggcacc cccgcccctt gtaaacttcc ctgaccctga
     1561 catgacaaga gttactaaca gcccctctct ccaagctcac ttacaggctc tctacttagt
     1621 ccagcacgaa gtctggagac ctctggcggc acgtaccaag aacaactgga ccgaccggtg
     1681 gtacctcacc cttaccgagt cggcgacaca gtgtgggtcc gccgacacca gactaagaac
     1741 ctagaacctc gctggaaagg accttacaca gtcctgctga ccacccccac cgccctcaaa
     1801 gtagacggca tcgcagcttg gatacacgcc gcccacgtga aggctgccga ccccgggggt
     1861 ggaccatcct ctagactgcc ggatccgcca ccatgatcag tctgattgcg gcgttagcgg
     1921 tagatcacgt tatcggcatg gaaaacgcca tgccgtggag cctgcctgcc gatctcgcct
     1981 ggtttaaacg caacacctta aataaacccg tgattatggg ccgccatacc tgggaatcaa
     2041 tcggtcgtcc gttgccagga cgcaaaaaca ttatcctcag cagtcaaccg agtacggacg
     2101 atcgcgtaac gtgggtgaag tcggcggatg aagccatcgc ggcgtgtggt gacgtaccag
     2161 aaatcatggt gattggcggc ggtcgcgttt atgagcagtt cttgccaaaa gcgcaaaaac
     2221 tgtatctgac gcatatcgac gcagaagtgg gaggcgacac ccatttcccg gattacgagc
     2281 cggatgactg gggatcggta ttcagcgaat tccacgatgc tgatgcgcag aactctcaca
     2341 gctattgctt tgtgattctg gggcggcgag catgcgtgag caagggcgag gagctgttca
     2401 ccggggtggt gcccatcctg gtcgagctgg acggcgacgt aaacggccac aagttcagcg
     2461 tgtccggcga gggcgagggc gatgccacct acggcaagct gaccctgaag ttcatctgca
     2521 ccaccggcaa gctgcccgtg ccctggccca ccctcgtgac caccttcggc tacggcctgc
     2581 agtgcttcgc ccgctacccc gaccacatga agcagcacga cttcttcaag tccgccatgc
     2641 ccgaaggcta cgtccaggag cgcaccatct tcttcaagga cgacggcaac tacaagaccc
     2701 gcgccgaggt gaagttcgag ggcgacaccc tggtgaaccg catcgagctg aagggcatcg
     2761 acttcaagga ggacggcaac atcctggggc acaagctgga gtacaactac aacagccaca
     2821 acgtctatat catggccgac aagcagaaga acggcatcaa ggtgaacttc aagatccgcc
     2881 acaacatcga ggacggcagc gtgcagctcg ccgaccacta ccagcagaac acccccatcg
     2941 gcgacggccc cgtgctgctg cccgacaacc actacctgag ctaccagtcc gccctgagca
     3001 aagaccccaa cgagaagcgc gatcacatgg tcctgctgga gttcgtgacc gccgccggga
     3061 tcactctcgg catggacgag ctgtacaagg tcgactatcc gtacgacgta ccagactacg
     3121 caccgcggcc gctacgtaaa ttccgcccct ctccctcccc cccccctaac gttactggcc
     3181 gaagccgctt ggaataaggc cggtgtgcgt ttgtctatat gttattttcc accatattgc
     3241 cgtcttttgg caatgtgagg gcccggaaac ctggccctgt cttcttgacg agcattccta
     3301 ggggtctttc ccctctcgcc aaaggaatgc aaggtctgtt gaatgtcgtg aaggaagcag
     3361 ttcctctgga agcttcttga agacaaacaa cgtctgtagc gaccctttgc aggcagcgga
     3421 accccccacc tggcgacagg tgcctctgcg gccaaaagcc acgtgtataa gatacacctg
     3481 caaaggcggc acaaccccag tgccacgttg tgagttggat agttgtggaa agagtcaaat
     3541 ggctctcctc aagcgtattc aacaaggggc tgaaggatgc ccagaaggta ccccattgta
     3601 tgggatctga tctggggcct cggtgcacat gctttacatg tgtttagtcg aggttaaaaa
     3661 aacgtctagg ccccccgaac cacggggacg tggttttcct ttgaaaaaca cgatgataat
     3721 atggccacaa ccatggtgag cggcctgctg aaggagagta tgcgcatcaa gatgtacatg
     3781 gagggcaccg tgaacggcca ctacttcaag tgcgagggcg agggcgacgg caaccccttc
     3841 gccggcaccc agagcatgag aatccacgtg accgagggcg cccccctgcc cttcgccttc
     3901 gacatcctgg ccccctgctg cgagtacggc agcaggacct tcgtgcacca caccgccgag
     3961 atccccgact tcttcaagca gagcttcccc gagggcttca cctgggagag aaccaccacc
     4021 tacgaggacg gcggcatcct gaccgcccac caggacacca gcctggaggg caactgcctg
     4081 atctacaagg tgaaggtgca cggcaccaac ttccccgccg acggccccgt gatgaagaac
     4141 aagagcggcg gctgggagcc cagcaccgag gtggtgtacc ccgagaacgg cgtgctgtgc
     4201 ggccggaacg tgatggccct gaaggtgggc gaccggcacc tgatctgcca ccactacacc
     4261 agctaccgga gcaagaaggc cgtgcgcgcc ctgaccatgc ccggcttcca cttcaccgac
     4321 atccggctcc agatgctgcg gaagaagaag gacgagtact tcgagctgta cgaggccagc
     4381 gtggcccggt acagcgacct gcccgagaag gccaacagat ctcccgggat ggtgagcggc
     4441 ctgctgaagg agagtatgcg catcaagatg tacatggagg gcaccgtgaa cggccactac
     4501 ttcaagtgcg agggcgaggg cgacggcaac cccttcgccg gcacccagag catgagaatc
     4561 cacgtgaccg agggcgcccc cctgcccttc gccttcgaca tcctggcccc ctgctgcgag
     4621 tacggcagca ggaccttcgt gcaccacacc gccgagatcc ccgacttctt caagcagagc
     4681 ttccccgagg gcttcacctg ggagagaacc accacctacg aggacggcgg catcctgacc
     4741 gcccaccagg acaccagcct ggagggcaac tgcctgatct acaaggtgaa ggtgcacggc
     4801 accaacttcc ccgccgacgg ccccgtgatg aagaacaaga gcggcggctg ggagcccagc
     4861 accgaggtgg tgtaccccga gaacggcgtg ctgtgcggcc ggaacgtgat ggccctgaag
     4921 gtgggcgacc ggcacctgat ctgccaccac tacaccagct accggagcaa gaaggccgtg
     4981 cgcgccctga ccatgcccgg cttccacttc accgacatcc ggctccagat gctgcggaag
     5041 aagaaggacg agtacttcga gctgtacgag gccagcgtgg cccggtacag cgacctgccc
     5101 gagaaggcca actgagtcga cgataaaata aaagatttta tttagtctcc agaaaaaggg
     5161 gggaatgaaa gaccccacct gtaggtttgg caagctagct taagtaacgc cattttgcaa
     5221 ggcatggaaa aatacataac tgagaataga gaagttcaga tcaaggtcag gaacagatgg
     5281 aacagctgaa tatgggccaa acaggatatc tgtggtaagc agttcctgcc ccggctcagg
     5341 gccaagaaca gatggaacag ctgaatatgg gccaaacagg atatctgtgg taagcagttc
     5401 ctgccccggc tcagggccaa gaacagatgg tccccagatg cggtccagcc ctcagcagtt
     5461 tctagagaac catcagatgt ttccagggtg ccccaaggac ctgaaatgac cctgtgcctt
     5521 atttgaacta accaatcagt tcgcttctcg cttctgttcg cgcgcttctg ctccccgagc
     5581 tcaataaaag agcccacaac ccctcactcg gggcgccagt cctccgattg actgagtcgc
     5641 ccgggtaccc gtgtatccaa taaaccctct tgcagttgca tccgacttgt ggtctcgctg
     5701 ttccttggga gggtctcctc tgagtgattg actacccgtc agcgggggtc tttcattcac
     5761 atgcagcatg tatcaaaatt aatttggttt tttttcttaa gtatttacat taaatggcca
     5821 tagttgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tggcgctctt
     5881 ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag
     5941 ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca
     6001 tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt
     6061 tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc
     6121 gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct
     6181 ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg
     6241 tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca
     6301 agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact
     6361 atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta
     6421 acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta
     6481 actacggcta cactagaagg acagtatttg gtatctgcgc tctgctgaag ccagttacct
     6541 tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt
     6601 tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga
     6661 tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca
     6721 tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agtttgcgca
     6781 aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg
     6841 aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg
     6901 tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc
     6961 gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg
     7021 agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg
     7081 aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag
     7141 gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat
     7201 caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc
     7261 cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc
     7321 ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa
     7381 ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaacac
     7441 gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt
     7501 cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc
     7561 gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa
     7621 caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca
     7681 tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc atgacattaa
     7741 cctataaaaa taggcgtatc acgaggccct ttcgtcttca agaacagctt tgctcttagg
     7801 agtttcctaa tacatcccaa actcaaatat ataaagcatt tgacttgttc tatgccctag
     7861 ggggcggggg gaagctaagc cagctttttt taacatttaa aatgttaatt ccattttaaa
     7921 tgcacagatg tttttatttc ataagggttt caatgtgcat gaatgctgca atattcctgt
     7981 taccaaagct agtataaata aaaatagata aacgtggaaa ttacttagag tttctgtcat
     8041 taacgtttcc ttcctcagtt gacaacataa atgcgctgct gagaagccag tttgcatctg
     8101 tcaggatcaa tttcccatta tgccagtcat attaattact agtcaattag ttgattttta
     8161 tttttgacat atacatg
//
