LOCUS       pENTR207-PRS-hRIP1-D138N(KD+RHIM)-no_stop      5113 bp    DNA     circular UNA 10-FEB-2012
FEATURES             Location/Qualifiers
     CDS             1..1716
                     /note="Region on donor sequence (hRIP1D138N -DD fragment +
                     primers): 14..1783"
                     /label="hRIP1D138N(KD + RHIM)"
     Misc._feature   complement(1721..1816)
                     /label=attL2
     Unsure          1728
                     /Design_Description="If necessary, linearize attB sequence
                     hRIP1D138N -DD fragment + primers at a unique REN site
                     outside the attB region. Ethanol precipitate and
                     resuspend.	Mix linearized attB sequence hRIP1D138N -DD
                     fragment + primers and pDONR Vector pDONR207 in BP buffer
                     and TE buffers.	Thaw, vortex and add BP Clonase enzyme.
                     Incubate >= 1 hr. at 25°.	Add Proteinase K for 10 min at
                     37°.	Transform into DH5a or TOP10 E. coli or any non-F'
                     episome bearing strain.	Select Entry Clones on agar plates
                     with antibiotic gentamicin."
                     /label="Gateway Cloning Description"
     Insertion       1729..5071
                     /note="Region on donor sequence (pDONR207): 2657..414"
                     /label=pDONR207
     Primer          complement(1841..1864)
                     /invitrogen="1994000"
                     /label="pDONR?E07 reverse primer"
     CDS             complement(2600..3133)
                     /invitrogen="1150000"
                     /label=Gm(R)
     Promoter        complement(3322..3349)
                     /invitrogen="2250000"
                     /label="Pc promoter"
     Terminator      4730..4757
                     /invitrogen="2590000"
                     /label="rrnB T2 transcription terminator"
     Terminator      4889..4932
                     /invitrogen="2580000"
                     /label="rrnB T1 transcription terminator"
     Primer          4957..4981
                     /invitrogen="1993000"
                     /label="pDONR?E07 forward primer"
     Misc._feature   4989..5084
                     /label=attL1
     Cleavage_site   5090..5113
                     /label="prescission cleavage site ( PRS)"
ORIGIN
        1 ATGCAACCAG ACATGTCCTT GAATGTCATT AAGATGAAAT CCAGTGACTT CCTGGAGAGT
       61 GCAGAACTGG ACAGCGGAGG CTTTGGGAAG GTGTCTCTGT GTTTCCACAG AACCCAGGGA
      121 CTCATGATCA TGAAAACAGT GTACAAGGGG CCCAACTGCA TTGAGCACAA CGAGGCCCTC
      181 TTGGAGGAGG CGAAGATGAT GAACAGACTG AGACACAGCC GGGTGGTGAA GCTCCTGGGC
      241 GTCATCATAG AGGAAGGGAA GTACTCCCTG GTGATGGAGT ACATGGAGAA GGGCAACCTG
      301 ATGCACGTGC TGAAAGCCGA GATGAGTACT CCGCTTTCTG TAAAAGGAAG GATAATTTTG
      361 GAAATCATTG AAGGAATGTG CTACTTACAT GGAAAAGGCG TGATACACAA GAACCTGAAG
      421 CCTGAAAATA TCCTTGTTGA TAATGACTTC CACATTAAGA TCGCAGACCT CGGCCTTGCC
      481 TCCTTTAAGA TGTGGAGCAA ACTGAATAAT GAAGAGCACA ATGAGCTGAG GGAAGTGGAC
      541 GGCACCGCTA AGAAGAATGG CGGCACCCTC TACTACATGG CGCCCGAGCA CCTGAATGAC
      601 GTCAACGCAA AGCCCACAGA GAAGTCGGAT GTGTACAGCT TTGCTGTAGT ACTCTGGGCG
      661 ATATTTGCAA ATAAGGAGCC ATATGAAAAT GCTATCTGTG AGCAGCAGTT GATAATGTGC
      721 ATAAAATCTG GGAACAGGCC AGATGTGGAT GACATCACTG AGTACTGCCC AAGAGAAATT
      781 ATCAGTCTCA TGAAGCTCTG CTGGGAAGCG AATCCGGAAG CTCGGCCGAC ATTTCCTGGC
      841 ATTGAAGAAA AATTTAGGCC TTTTTATTTA AGTCAATTAG AAGAAAGTGT AGAAGAGGAC
      901 GTGAAGAGTT TAAAGAAAGA GTATTCAAAC GAAAATGCAG TTGTGAAGAG AATGCAGTCT
      961 CTTCAACTTG ATTGTGTGGC AGTACCTTCA AGCCGGTCAA ATTCAGCCAC AGAACAGCCT
     1021 GGTTCACTGC ACAGTTCCCA GGGACTTGGG ATGGGTCCTG TGGAGGAGTC CTGGTTTGCT
     1081 CCTTCCCTGG AGCACCCACA AGAAGAGAAT GAGCCCAGCC TGCAGAGTAA ACTCCAAGAC
     1141 GAAGCCAACT ACCATCTTTA TGGCAGCCGC ATGGACAGGC AGACGAAACA GCAGCCCAGA
     1201 CAGAATGTGG CTTACAACAG AGAGGAGGAA AGGAGACGCA GGGTCTCCCA TGACCCTTTT
     1261 GCACAGCAAA GACCTTACGA GAATTTTCAG AATACAGAGG GAAAAGGCAC TGCTTATTCC
     1321 AGTGCAGCCA GTCATGGTAA TGCAGTGCAC CAGCCCTCAG GGCTCACCAG CCAACCTCAA
     1381 GTACTGTATC AGAACAATGG ATTATATAGC TCACATGGCT TTGGAACAAG ACCACTGGAT
     1441 CCAGGAACAG CAGGTCCCAG AGTTTGGTAC AGGCCAATTC CAAGTCATAT GCCTAGTCTG
     1501 CATAATATCC CAGTGCCTGA GACCAACTAT CTAGGAAATA CACCCACCAT GCCATTCAGC
     1561 TCCTTGCCAC CAACAGATGA ATCTATAAAA TATACCATAT ACAATAGTAC TGGCATTCAG
     1621 ATTGGAGCCT ACAATTATAT GGAGATTGGT GGGACGAGTT CATCACTACT AGACAGCACA
     1681 AATACGAACT TCAAAGAAGA GCCAGCTGCT AAGTACTACC CAGCTTTCTT GTACAAAGTG
     1741 GGCATTATAA GAAAGCATTG CTTATCAATT TGTTGCAACG AACAGGTCAC TATCAGTCAA
     1801 AATAAAATCA TTATTTGCCA TCCAGCTGCA GCTCTGGCCC GTGTCTCAAA ATCTCTGATG
     1861 TTACATTGCA CAAGATAAAA ATATATCATC ATGAACAATA AAACTGTCTG CTTACATAAA
     1921 CAGTAATACA AGGGGTGTTA TGAGCCATAT TCAACGGGAA ACGTCGAGGC CGCGATTAAA
     1981 TTCCAACATG GATGCTGATT TATATGGGTA TAAATGGGCT CGCGATAATG TCGGGCAATC
     2041 AGGTGCGACA ATCTATCGCT TGTATGGGAA GCCCGATGCG CCAGAGTTGT TTCTGAAACA
     2101 TGGCAAAGGT AGCGTTGCCA ATGATGTTAC AGATGAGATG GTCAGACTAA ACTGGCTGAC
     2161 GGAATTTATG CCTCTTCCGA CCATCAAGCA TTTTATCCGT ACTCCTGATG ATGCATGGTT
     2221 ACTCACCACT GCGATCCCCG GAAAAACAGC ATTCCAGGTA TTAGAAGAAT ATCCTGATTC
     2281 AGGTGAAAAT ATTGTTGATG CGCTGGCAGT GTTCCTGCGC CGGTTGCATT CGATTCCTGT
     2341 TTGTAATTGT CCTTTTAACA GCGATCGCGT ATTTCGTCTC GCTCAGGCGC AATCACGAAT
     2401 GAATAACGGT TTGGTTGATG CGAGTGATTT TGATGACGAG CGTAATGGCT GGCCTGTTGA
     2461 ACAAGTCTGG AAAGAAATGC ATAAACTTTT GCCATTCTCA CCGGATTCAG TCGTCACTCA
     2521 TGGTGATTTC TCACTTGATA ACCTTATTTT TGACGAGGGG AAATTAGGCC GGGAAGCCGA
     2581 TCTCGGCTTG AACGAATTGT TAGGTGGCGG TACTTGGGTC GATATCAAAG TGCATCACTT
     2641 CTTCCCGTAT GCCCAACTTT GTATAGAGAG CCACTGCGGG ATCGTCACCG TAATCTGCTT
     2701 GCACGTAGAT CACATAAGCA CCAAGCGCGT TGGCCTCATG CTTGAGGAGA TTGATGAGCG
     2761 CGGTGGCAAT GCCCTGCCTC CGGTGCTCGC CGGAGACTGC GAGATCATAG ATATAGATCT
     2821 CACTACGCGG CTGCTCAAAC CTGGGCAGAA CGTAAGCCGC GAGAGCGCCA ACAACCGCTT
     2881 CTTGGTCGAA GGCAGCAAGC GCGATGAATG TCTTACTACG GAGCAAGTTC CCGAGGTAAT
     2941 CGGAGTCCGG CTGATGTTGG GAGTAGGTGG CTACGTCTCC GAACTCACGA CCGAAAAGAT
     3001 CAAGAGCAGC CCGCATGGAT TTGACTTGGT CAGGGCCGAG CCTACATGTG CGAATGATGC
     3061 CCATACTTGA GCCACCTAAC TTTGTTTTAG GGCGACTGCC CTGCTGCGTA ACATCGTTGC
     3121 TGCTGCGTAA CATCGTTGCT GCTCCATAAC ATCAAACATC GACCCACGGC GTAACGCGCT
     3181 TGCTGCTTGG ATGCCCGAGG CATAGACTGT ACAAAAAAAC AGTCATAACA AGCCATGAAA
     3241 ACCGCCACTG CGCCGTTACC ACCGCTGCGT TCGGTCAAGG TTCTGGACCA GTTGCGTGAG
     3301 CGCATACGCT ACTTGCATTA CAGTTTACGA ACCGAACAGG CTTATGTCAA CTGGGTTCGT
     3361 GCCTTCATCC GTTTCCACGG TGTGCGTCAC CCGGCAACCT TGGGCAGCAG CGAAGTCGAG
     3421 GCATTTCTGT CCTGGCTGGC GAACGAGCGC AAGGTTTCGG TCTCCACGCA TCGTCAGGCA
     3481 TTGGCGGCCT TGCTGTTCTT CTACGGCAAG GTGCTGTGCA CGGATCTGCC CTGGCTTCAG
     3541 GAGATCGGAA GACCTCGGCC GTCGCGGCGC TTGCCGGTGG TGCTGACCCC GGATGAAGTG
     3601 GTTCGCATCC TCGGTTTTCT GGAAGGCGAG CATCGTTTGT TCGCCCAGGA CTCTAGCTAT
     3661 AGTTCTAGTG GTTGGCTACT AATAGGTTGT ATTGATGTTG GACGAGTCGG AATCGCAGAC
     3721 CGATACCAGG ATCTTGCCAT CCTATGGAAC TGCCTCGGTG AGTTTTCTCC TTCATTACAG
     3781 AAACGGCTTT TTCAAAAATA TGGTATTGAT AATCCTGATA TGAATAAATT GCAGTTTCAT
     3841 TTGATGCTCG ATGAGTTTTT CTAATCAGAA TTGGTTAATT GGTTGTAACA CTGGCAGAGC
     3901 ATTACGCTGA CTTGACGGGA CGGCGCAAGC TCATGACCAA AATCCCTTAA CGTGAGTTTT
     3961 CGTTCCACTG AGCGTCAGAC CCCGTAGAAA AGATCAAAGG ATCTTCTTGA GATCCTTTTT
     4021 TTCTGCGCGT AATCTGCTGC TTGCAAACAA AAAAACCACC GCTACCAGCG GTGGTTTGTT
     4081 TGCCGGATCA AGAGCTACCA ACTCTTTTTC CGAAGGTAAC TGGCTTCAGC AGAGCGCAGA
     4141 TACCAAATAC TGTCCTTCTA GTGTAGCCGT AGTTAGGCCA CCACTTCAAG AACTCTGTAG
     4201 CACCGCCTAC ATACCTCGCT CTGCTAATCC TGTTACCAGT GGCTGCTGCC AGTGGCGATA
     4261 AGTCGTGTCT TACCGGGTTG GACTCAAGAC GATAGTTACC GGATAAGGCG CAGCGGTCGG
     4321 GCTGAACGGG GGGTTCGTGC ACACAGCCCA GCTTGGAGCG AACGACCTAC ACCGAACTGA
     4381 GATACCTACA GCGTGAGCTA TGAGAAAGCG CCACGCTTCC CGAAGGGAGA AAGGCGGACA
     4441 GGTATCCGGT AAGCGGCAGG GTCGGAACAG GAGAGCGCAC GAGGGAGCTT CCAGGGGGAA
     4501 ACGCCTGGTA TCTTTATAGT CCTGTCGGGT TTCGCCACCT CTGACTTGAG CGTCGATTTT
     4561 TGTGATGCTC GTCAGGGGGG CGGAGCCTAT GGAAAAACGC CAGCAACGCG GCCTTTTTAC
     4621 GGTTCCTGGC CTTTTGCTGG CCTTTTGCTC ACATGTTCTT TCCTGCGTTA TCCCCTGATT
     4681 CTGTGGATAA CCGTATTACC GCTAGCCAGG AAGAGTTTGT AGAAACGCAA AAAGGCCATC
     4741 CGTCAGGATG GCCTTCTGCT TAGTTTGATG CCTGGCAGTT TATGGCGGGC GTCCTGCCCG
     4801 CCACCCTCCG GGCCGTTGCT TCACAACGTT CAAATCCGCT CCCGGCGGAT TTGTCCTACT
     4861 CAGGAGAGCG TTCACCGACA AACAACAGAT AAAACGAAAG GCCCAGTCTT CCGACTGAGC
     4921 CTTTCGTTTT ATTTGATGCC TGGCAGTTCC CTACTCTCGC GTTAACGCTA GCATGGATCT
     4981 CGGGCCCCAA ATAATGATTT TATTTTGACT GATAGTGACC TGTTCGTTGC AACAAATTGA
     5041 TGAGCAATGC TTTTTTATAA TGCCAAGTTT GTACAAAAAA GCAGGCTTAC TGGAAGTTCT
     5101 GTTCCAGGGG CCC
//
