LOCUS       Exported                8176 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  ecDHFR based destable domain function at low temperature for 
            regulating protein stability in Caenorhabditis elegans.
ACCESSION   .
VERSION     .
KEYWORDS    pBMN YFP-ltDHFR(DD)
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8176)
  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 8176)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..8176
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     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             1896..1922
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             1923..2639
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1968..1989)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2229..2248)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2576..2597
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             2646..3122
                     /codon_start=1
                     /gene="folA"
                     /product="E. coli dihydrofolate reductase"
                     /label=ecDHFR
                     /translation="ISLIAALAVDHVIGMENAMPWNLPADLAWFKRNTLDKPVIMGRHT
                     WESIGRPLPGRKDIILSSQPSTDDRVTWVKSVDEAIAACGDVPEIMVIGGGRVYEQFLP
                     KAQKLYLTHIDAEVEGDTHFPDYEPDGWESVFSEFHDADAQNSHSYCFEILERR"
     misc_feature    3146..3731
                     /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             3732..4415
                     /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             4428..5114
                     /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             5163..5756
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     primer_bind     complement(5887..5904)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6058..6646)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6138..6157)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(6820..7680)
                     /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     7443..7462
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     primer_bind     7748..7766
                     /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 ccatgtatcc gtacgacgta ccggattatg
     1921 cgatggtgag caagggcgag gagctgttca ccggggtggt gcccatcctg gtcgagctgg
     1981 acggcgacgt aaacggccac aagttcagcg tgtccggcga gggcgagggc gatgccacct
     2041 acggcaagct gaccctgaag ttcatctgca ccaccggcaa gctgcccgtg ccctggccca
     2101 ccctcgtgac caccttcggc tacggcctgc agtgcttcgc ccgctacccc gaccacatga
     2161 agcagcacga cttcttcaag tccgccatgc ccgaaggcta cgtccaggag cgcaccatct
     2221 tcttcaagga cgacggcaac tacaagaccc gcgccgaggt gaagttcgag ggcgacaccc
     2281 tggtgaaccg catcgagctg aagggcatcg acttcaagga ggacggcaac atcctggggc
     2341 acaagctgga gtacaactac aacagccaca acgtctatat catggccgac aagcagaaga
     2401 acggcatcaa ggtgaacttc aagatccgcc acaacatcga ggacggcagc gtgcagctcg
     2461 ccgaccacta ccagcagaac acccccatcg gcgacggccc cgtgctgctg cccgacaacc
     2521 actacctgag ctaccagtcc gccctgagca aagaccccaa cgagaagcgc gatcacatgg
     2581 tcctgctgga gttcgtgacc gccgccggga tcactctcgg catggacgag ctgtacaagc
     2641 tcgagatcag tctgattgcg gcgttagcgg tagatcacgt tatcggcatg gaaaacgcca
     2701 tgccgtggaa cctgcctgcc gatctcgcct ggtttaaacg caacacctta gataaacccg
     2761 tgattatggg ccgccatacc tgggaatcaa tcggtcgtcc gttgccagga cgcaaagata
     2821 ttatcctcag cagtcaaccg agtacggacg atcgcgtaac gtgggtgaag tcggtggatg
     2881 aagccatcgc ggcgtgtggt gacgtaccag aaatcatggt gattggcggc ggtcgcgttt
     2941 atgaacagtt cttgccaaaa gcgcaaaaac tgtatctgac gcatatcgac gcagaagtgg
     3001 aaggcgacac ccatttcccg gattacgagc cggatggctg ggaatcggta ttcagcgaat
     3061 tccacgatgc tgatgcgcag aactctcaca gctattgctt tgagattctg gagcggcgat
     3121 gaccgcggcc 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 acgtctaggc cccccgaacc acggggacgt ggttttcctt tgaaaaacac gatgataata
     3721 tggccacaac catggtgagc ggcctgctga aggagagtat gcgcatcaag atgtacatgg
     3781 agggcaccgt gaacggccac tacttcaagt gcgagggcga gggcgacggc aaccccttcg
     3841 ccggcaccca gagcatgaga atccacgtga ccgagggcgc ccccctgccc ttcgccttcg
     3901 acatcctggc cccctgctgc gagtacggca gcaggacctt cgtgcaccac accgccgaga
     3961 tccccgactt cttcaagcag agcttccccg agggcttcac ctgggagaga accaccacct
     4021 acgaggacgg cggcatcctg accgcccacc aggacaccag cctggagggc aactgcctga
     4081 tctacaaggt gaaggtgcac ggcaccaact tccccgccga cggccccgtg atgaagaaca
     4141 agagcggcgg ctgggagccc agcaccgagg tggtgtaccc cgagaacggc gtgctgtgcg
     4201 gccggaacgt gatggccctg aaggtgggcg accggcacct gatctgccac cactacacca
     4261 gctaccggag caagaaggcc gtgcgcgccc tgaccatgcc cggcttccac ttcaccgaca
     4321 tccggctcca gatgctgcgg aagaagaagg acgagtactt cgagctgtac gaggccagcg
     4381 tggcccggta cagcgacctg cccgagaagg ccaacagatc tcccgggatg gtgagcggcc
     4441 tgctgaagga gagtatgcgc atcaagatgt acatggaggg caccgtgaac ggccactact
     4501 tcaagtgcga gggcgagggc gacggcaacc ccttcgccgg cacccagagc atgagaatcc
     4561 acgtgaccga gggcgccccc ctgcccttcg ccttcgacat cctggccccc tgctgcgagt
     4621 acggcagcag gaccttcgtg caccacaccg ccgagatccc cgacttcttc aagcagagct
     4681 tccccgaggg cttcacctgg gagagaacca ccacctacga ggacggcggc atcctgaccg
     4741 cccaccagga caccagcctg gagggcaact gcctgatcta caaggtgaag gtgcacggca
     4801 ccaacttccc cgccgacggc cccgtgatga agaacaagag cggcggctgg gagcccagca
     4861 ccgaggtggt gtaccccgag aacggcgtgc tgtgcggccg gaacgtgatg gccctgaagg
     4921 tgggcgaccg gcacctgatc tgccaccact acaccagcta ccggagcaag aaggccgtgc
     4981 gcgccctgac catgcccggc ttccacttca ccgacatccg gctccagatg ctgcggaaga
     5041 agaaggacga gtacttcgag ctgtacgagg ccagcgtggc ccggtacagc gacctgcccg
     5101 agaaggccaa ctgagtcgac gataaaataa aagattttat ttagtctcca gaaaaagggg
     5161 ggaatgaaag accccacctg taggtttggc aagctagctt aagtaacgcc attttgcaag
     5221 gcatggaaaa atacataact gagaatagag aagttcagat caaggtcagg aacagatgga
     5281 acagctgaat atgggccaaa caggatatct gtggtaagca gttcctgccc cggctcaggg
     5341 ccaagaacag atggaacagc tgaatatggg ccaaacagga tatctgtggt aagcagttcc
     5401 tgccccggct cagggccaag aacagatggt ccccagatgc ggtccagccc tcagcagttt
     5461 ctagagaacc atcagatgtt tccagggtgc cccaaggacc tgaaatgacc ctgtgcctta
     5521 tttgaactaa ccaatcagtt cgcttctcgc ttctgttcgc gcgcttctgc tccccgagct
     5581 caataaaaga gcccacaacc cctcactcgg ggcgccagtc ctccgattga ctgagtcgcc
     5641 cgggtacccg tgtatccaat aaaccctctt gcagttgcat ccgacttgtg gtctcgctgt
     5701 tccttgggag ggtctcctct gagtgattga ctacccgtca gcgggggtct ttcattcaca
     5761 tgcagcatgt atcaaaatta atttggtttt ttttcttaag tatttacatt aaatggccat
     5821 agttgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt ggcgctcttc
     5881 cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc
     5941 tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat
     6001 gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt
     6061 ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg
     6121 aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc
     6181 tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt
     6241 ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa
     6301 gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta
     6361 tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa
     6421 caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa
     6481 ctacggctac actagaagga cagtatttgg tatctgcgct ctgctgaagc cagttacctt
     6541 cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt
     6601 ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat
     6661 cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat
     6721 gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa gtttgcgcaa
     6781 atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga
     6841 ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt
     6901 gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg
     6961 agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga
     7021 gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga
     7081 agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg
     7141 catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc
     7201 aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc
     7261 gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca
     7321 taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac
     7381 caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaacacg
     7441 ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc
     7501 ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg
     7561 tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac
     7621 aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat
     7681 actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca tgacattaac
     7741 ctataaaaat aggcgtatca cgaggccctt tcgtcttcaa gaacagcttt gctcttagga
     7801 gtttcctaat acatcccaaa ctcaaatata taaagcattt gacttgttct atgccctagg
     7861 gggcgggggg aagctaagcc agcttttttt aacatttaaa atgttaattc cattttaaat
     7921 gcacagatgt ttttatttca taagggtttc aatgtgcatg aatgctgcaa tattcctgtt
     7981 accaaagcta gtataaataa aaatagataa acgtggaaat tacttagagt ttctgtcatt
     8041 aacgtttcct tcctcagttg acaacataaa tgcgctgctg agaagccagt ttgcatctgt
     8101 caggatcaat ttcccattat gccagtcata ttaattacta gtcaattagt tgatttttat
     8161 ttttgacata tacatg
//
