LOCUS       40924_33727        5785 bp DNA     circular SYN 01-JAN-1980
DEFINITION  Vector for constitutive expression of a thermolabile Flp recombinase
            (flp-F70L) in mammalian cells, for use with the Flp-In(TM) system.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5785)
  AUTHORS   Ben Glick
  TITLE     Direct Submission
REFERENCE   2  (bases 1 to 5785)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"
COMMENT     This plasmid does not contain a drug resistance marker, and will be 
            gradually lost during cell culture.
FEATURES             Location/Qualifiers
     source          1..5785
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        237..616
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        617..820
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early
                     promoter"
     promoter        865..883
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     intron          932..1161
                     /label=chimeric intron
                     /note="chimera between introns from adenovirus and 
                     immunoglobulin heavy chain genes"
     CDS             1202..2470
                     /codon_start=1
                     /label=FLP
                     /note="site-specific recombinase"
                     /translation="MPQFDILCKTPPKVLVRQFVERFERPSGEKIALCAAELTYLCWMI
                     THNGTAIKRATFMSYNTIISNSLSLDIVNKSLQFKYKTQKATILEASLKKLIPAWEFTI
                     IPYYGQKHQSDITDIVSSLQLQFESSEEADKGNSHSKKMLKALLSEGESIWEITEKILN
                     SFEYTSRFTKTKTLYQFLFLATFINCGRFSDIKNVDPKSFKLVQNKYLGVIIQCLVTET
                     KTSVSRHIYFFSARGRIDPLVYLDEFLRNSEPVLKRVNRTGNSSSNKQEYQLLKDNLVR
                     SYNKALKKNAPYSIFAIKNGPKSHIGRHLMTSFLSMKGLTELTNVVGNWSDKRASAVAR
                     TTYTHQITAIPDHYFALVSRYYAYDPISKEMIALKDETNPIEEWQHIEQLKGSAEGSIR
                     YPAWNGIISQEVLDYLSSYINRRI"
     polyA_signal    complement(2610..2731)
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     promoter        complement(2945..2963)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2984..3000)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(3008..3024)
                     /label=lac operator
                     /note="The lac repressor binds to the lac operator to
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(3032..3062)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    complement(3077..3098)
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     rep_origin      complement(3386..3967)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(4141..4998)
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(4999..5103)
                     /label=AmpR promoter
     rep_origin      complement(5128..5583)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     primer_bind     5725..5741
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 ccgcggtctt tcggactttt gaaagtgatg gtggtggggg aaggattcga accttcgaag
       61 tcgatgacgg cagatttaga gtctgctccc tgcttgtgtg ttggaggtcg ctgagtagtg
      121 cgcgagcaaa atttaagcta caacaaggca aggcttgacc gacaattgca tgaagaatct
      181 gcttagggtt aggcgttttg cgctgcttcg cgatgtacgg gccagatata cgcgttgaca
      241 ttgattattg actagttatt aatagtaatc aattacgggg tcattagttc atagcccata
      301 tatggagttc cgcgttacat aacttacggt aaatggcccg cctggctgac cgcccaacga
      361 cccccgccca ttgacgtcaa taatgacgta tgttcccata gtaacgccaa tagggacttt
      421 ccattgacgt caatgggtgg agtatttacg gtaaactgcc cacttggcag tacatcaagt
      481 gtatcatatg ccaagtacgc cccctattga cgtcaatgac ggtaaatggc ccgcctggca
      541 ttatgcccag tacatgacct tatgggactt tcctacttgg cagtacatct acgtattagt
      601 catcgctatt accatggtga tgcggttttg gcagtacatc aatgggcgtg gatagcggtt
      661 tgactcacgg ggatttccaa gtctccaccc cattgacgtc aatgggagtt tgttttggca
      721 ccaaaatcaa cgggactttc caaaatgtcg taacaactcc gccccattga cgcaaatggg
      781 cggtaggcgt gtacggtggg aggtctatat aagcagagct ctctggctaa ctagagaacc
      841 cactgcttac tggcttatcg aaattaatac gactcactat agggagaccc aagcttctag
      901 aattcgctgt ctgcgagggc cagctgttgg ggtgagtact ccctctcaaa agcgggcatg
      961 acttctgcgc taagattgtc agtttccaaa aacgaggagg atttgatatt cacctggccc
     1021 gcggtgatgc ctttgagggt ggccgcgtcc atctggtcag aaaagacaat ctttttgttg
     1081 tcaagcttga ggtgtggcag gcttgagatc tggccataca cttgagtgac aatgacatcc
     1141 actttgcctt tctctccaca ggtgtccact cccaggtcca actgcagccc aagcttccac
     1201 catgccacaa tttgatatat tatgtaaaac accacctaag gtgcttgttc gtcagtttgt
     1261 ggaaaggttt gaaagacctt caggtgagaa aatagcatta tgtgctgctg aactaaccta
     1321 tttatgttgg atgattacac ataacggaac agcaatcaag agagccacat tcatgagcta
     1381 taatactatc ataagcaatt cgctgagttt ggatattgtc aacaagtcac tgcagtttaa
     1441 atacaagacg caaaaagcaa caattctgga agcctcatta aagaaattga ttcctgcttg
     1501 ggaatttaca attattcctt actatggaca aaaacatcaa tctgatatca ctgatattgt
     1561 aagtagtttg caattacagt tcgaatcatc ggaagaagca gataagggaa atagccacag
     1621 taaaaaaatg cttaaagcac ttctaagtga gggtgaaagc atctgggaga tcactgagaa
     1681 aatactaaat tcgtttgagt atacttcgag atttacaaaa acaaaaactt tataccaatt
     1741 cctcttccta gctactttca tcaattgtgg aagattcagc gatattaaga acgttgatcc
     1801 gaaatcattt aaattagtcc aaaataagta tctgggagta ataatccagt gtttagtgac
     1861 agagacaaag acaagcgtta gtaggcacat atacttcttt agcgcaaggg gtaggatcga
     1921 tccacttgta tatttggatg aatttttgag gaattctgaa ccagtcctaa aacgagtaaa
     1981 taggaccggc aattcttcaa gcaacaagca ggaataccaa ttattaaaag ataacttagt
     2041 cagatcgtac aacaaagctt tgaagaaaaa tgcgccttat tcaatctttg ctataaaaaa
     2101 tggcccaaaa tctcacattg gaagacattt gatgacctca tttctttcaa tgaagggcct
     2161 aacggagttg actaatgttg tgggaaattg gagcgataag cgtgcttctg ccgtggccag
     2221 gacaacgtat actcatcaga taacagcaat acctgatcac tacttcgcac tagtttctcg
     2281 gtactatgca tatgatccaa tatcaaagga aatgatagca ttgaaggatg agactaatcc
     2341 aattgaggag tggcagcata tagaacagct aaagggtagt gctgaaggaa gcatacgata
     2401 ccccgcatgg aatgggataa tatcacagga ggtactagac tacctttcat cctacataaa
     2461 tagacgcata taagtacgca tttaagcata aacacgcact atgccgttct tctcatgtat
     2521 atatatatac aggcaacacg cagatatagg tgcgacgtga acagtgagct gtatgtgcgg
     2581 gtaccgagct cctcgaggat cagacatgat aagatacatt gatgagtttg gacaaaccac
     2641 aactagaatg cagtgaaaaa aatgctttat ttgtgaaatt tgtgatgcta ttgctttatt
     2701 tgtaaccatt ataagctgca ataaacaagt tatcctcgag gagctcatga gcgcttgttt
     2761 cggcgtgggt atggtggcag gcccgtggcc gggggactgt tgggcgccat ctccttgcat
     2821 gcaccattcc ttgcggcggc ggtgctcaac ggcctcaacc tactactggg ctgcttccta
     2881 atgcaggagt cgcataaggg agagcgtcga cctcgagggg gggcccggta cccagctttt
     2941 gttcccttta gtgagggtta attgcgcgct tggcgtaatc atggtcatag ctgtttcctg
     3001 tgtgaaattg ttatccgctc acaattccac acaacatacg agccgggagc ataaagtgta
     3061 aagcctgggg tgcctaatga gtgagctaac tcacattaat tgcgttgcgc tcactgcccg
     3121 ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga
     3181 gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg
     3241 tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag
     3301 aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc
     3361 gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca
     3421 aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt
     3481 ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc
     3541 tgtccgcctt tctcttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc
     3601 agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc
     3661 gaccctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat
     3721 cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta
     3781 cagagttctt gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct
     3841 gcgctctgct gaagccagtt acttcggaaa aagagttggt agctcttgat ccggcaaaca
     3901 aaccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg
     3961 atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc
     4021 acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa
     4081 ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta
     4141 ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt
     4201 tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag
     4261 tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag caataaacca
     4321 gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc
     4381 tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt
     4441 tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag
     4501 ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt
     4561 tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat
     4621 ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt
     4681 gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc
     4741 ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat
     4801 cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag
     4861 ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt
     4921 ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg
     4981 gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta
     5041 ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc
     5101 gcgcacattt ccccgaaaag tgccactaaa ttgtaagcgt taatattttg ttaaaattcg
     5161 cgttaaattt ttgttaaatc agctcatttt ttaaccaata ggccgaaatc ggcaaaatcc
     5221 cttataaatc aaaagaatag accgagatag ggttgagtgt tgttccagtt tggaacaaga
     5281 gtccactatt aaagaacgtg gactccaacg tcaaagggcg aaaaaccgtc tatcagggcg
     5341 atggcccact acgtgaacca tcaccctaat caagtttttt ggggtcgagg tgccgtaaag
     5401 cactaaatcg gaaccctaaa gggagccccc gatttagagc ttgacgggga aagccggcga
     5461 acgtggcgag aaaggaaggg aagaaagcga aaggagcggg cgctagggcg ctggcaagtg
     5521 tagcggtcac gctgcgcgta accaccacac ccgccgcgct taatgcgccg ctacagggcg
     5581 cgtcccattc gccattcagg ctgcgcaact gttgggaagg gcgatcggtg cgggcctctt
     5641 cgctattacg ccagctggcg aaagggggat gtgctgcaag gcgattaagt tgggtaacgc
     5701 cagggttttc ccagtcacga cgttgtaaaa cgacggccag tgagcgcgcg taatacgact
     5761 cactataggg cgaattggag ctcca
//
