Please review the QC test results indicated by icon below as well as clone information before placing your order.

pcDNA4c/His-Xpress-HA-hBIG2(E738K) (#RDB15365)

Expression vector of human BIG2, dominant-negative mutant E738K.

Alternative name	N8
Clone info.	Expression vector of human Arf guanine nucleotide-exchange factors 2 (BIG2), dominant-negative mutant E738K, tagged with Xpress and HA epitope at N-ter. This clone contains a cDNA fragment corresponding to the 180nt to 5972nt of NM_006420.3. Known differences (at least) : A to G substitution at 2390nt (Gly, synonym) ; A to G substitution at 2399nt (Gln, synonym) ; T to C substitution at 2405nt (Ile, synonym) ; C to T substitution at 2408nt (Asp, synonym) ; C to T substitution at 5342nt (Leu, synonym) ; A to G substitution at 5345nt (Lys, synonym).
Comment	1. BIG1 and BIG2 containing plasmids seem to be toxic to bacteria. After transformation, you need longer incubation time to get colonies as compared with usual transformed bacteria. Moreover, the colonies are smaller than usual ones. So, please take small colonies for liquid culture (The depositor is usually using DH5alpha for transformation). 2. Mini-prep: For liquid culture, you should use very fresh colonies otherwise you will get very low amounts of plasmids. 3. Maxi-prep (Large scale prep): Do not make a preculture from a single colony for large scale culture. You will get very low amounts of plasmids. The depositor would recommend you that after transformation and recovery culture, bring all transformed bacteria directly to large scale culture (200 ml or more of ampicillin containing medium). This helps a lot to get plasmids. Expression was confirmed by the depositor with immunostaining.
Vector backbone	pcDNA4/HisMax (plasmid)
Size of vector backbone	5.3 kb
Selectable markers	Amp^r (E. coli), Zeo^r (mammalian cells).
Gene/insert name	human BIG2 cDNA
Depositor|Developer	Nakayama, Kazuhisa |

 

Distribution information

Please check terms and conditions set forth by the depositor, which are specified in the RIKEN BRC Catalog and/or Web Catalog.

Ordering forms	Order form [Credit Card Payment] [Bank Transfer Payment] [Example of order form ] MTA, for use for not-for-profit academic purpose [Word] [Example of MTA ] Please visit Information of Request for Distribution.[link]
Terms and conditions for distribution	In publishing the research results obtained by use of the BIOLOGICAL RESOURCE, a citation of literature designated by the DEPOSITOR is requested (Ishizaki, R. et al. Mol. Biol. Cell, 19 (6): 2650-2660, 2008). RECIPIENT agrees to use the BIOLOGICAL RESOURCE only for academic research in the non-profit organization.

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Catalog #	Resource name	Shipping form	Fee (non-profit org.)
RDB15365	pcDNA4c/His-Xpress-HA-hBIG2(E738K)	DNA solution

Please review the QC test results indicated by icon below as well as clone information before placing your order.

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How to cite this biological resource

Materials & Methods section:

The pcDNA4c/His-Xpress-HA-hBIG2(E738K) was provided by the RIKEN BRC through the National BioResource Project of the MEXT, Japan (cat. RDB15365).

Reference section:

Ishizaki, R., Shin, H.W., Mitsuhashi, H., Nakayama, K., Redundant roles of BIG2 and BIG1, guanine-nucleotide exchange factors for ADP-ribosylation factors in membrane traffic between the trans-Golgi network and endosomes. Mol. Biol. Cell 19 (6): 2650-2660 (2008). PMID 18417613. [link to RRC of NBRP]

Further references such as user reports and related articles (go to bottom)

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QC test results

RIKEN BRC has sequenced portions of this material for quality test.
Please review the QC test results indicated by icon as well as clone information before placing your order.

Test sheet	RDB15365_A7I5p1-1.pdf

Nucleotide sequence of a portion of this resource (if available).

Primer: CMV_Forward (Pr0016) Region: CMV pro, QBI SP163, 6xHis, T7 tag, Xpress tag, HA, insert 5' Sequence file: RDB15365_A7I5a.seq
>D03588D2_A7I5_2_CMV_Forward_A09_03_ABI24.ab1     1 NNNNNNNNNN NNNNNTNNNN GCNNNNGGGC GGTANGCGTG TACGGTGGGN NGTCTATATA    61 AGCAGAGCTC TCTGGCTAAC TAGAGAACCC ACTGCTTACT GGCTTATCGA AATTAATACG   121 ACTCACTATA GGGAGACCCA AGCTGGCTAG CGTTTAAACT TAAGCTTAGC GCAGAGGCTT   181 GGGGCAGCCG AGCGGCAGCC AGGCCCCGGC CCGGGCCTCG GTTCCAGAAG GGAGAGGAGC   241 CCGCCAAGGC GCGCAAGAGA GCGGGCTGCC TCGCAGTCCG AGCCGGAGAG GGAGCGCGAG   301 CCGCGCCGGC CCCGGACGGC CTCCGAAACC ATGGGGGGTT CTCATCATCA TCATCATCAT   361 GGTATGGCTA GCATGACTGG TGGACAGCAA ATGGGTCGGG ATCTGTACGA CGATGACGAT   421 AAGGTACCTA CCATGTACCC GTACGACGTG CCCGACTACG CGGGATCCAC CATGCAGGAG   481 AGCCAGACCA AGAGCATGTT CGTGTCCCGG GCCCTGGAGA AGATCCTAGC CGACAAGGAG   541 GTGAAGCGGC CCCAGCACTC CCAGCTGCGC AGGGCCTGCC AGGTGGCGCT CGATGAAATT   601 AAAGCAGAAA TAGAAAAGCA GAGGCTTGGC ACTGCTGCAC CACCAAAGGC AAACTTCATT   661 GAAGCTGACA AGTATTTTCT TCCNTTCGAG CTAGCTTGCC AGTCCAAGTC CCCAAGGGTA   721 GTCAGCACAT CCCTTGACTG CTTGCAGAAA CTCATCGCAT ACGGGCACAT CACTGGCAAC   781 GCCCCTGACA GTGGANCCCC NGGGAAGCGG CTGANCGACA GAATTGTTGA AACCATTTGC   841 AGTTGTTTTC NGGNCCCTCN NANNGATGAA NGGGTTCAGT TACNAATNAT TNANNNNCNN   901 CNGNNNGCNN NGNNNTCCCN NCNCNNNNNA ANTNNNNNNG NNNNNATCNN NNNNANNNNN 961 NNNNN //
Primer: BGH_rev2 (Pr0606) Region: BGH pA, insert 3' Sequence file: RDB15365_A7I5b.seq
>D03588D2_A7I5_2_BGH_rev2_F05_06_ABI08.ab1     1 NNNNNNNNNN NNNNNNNNNN NNNNGNGNNN NNNCAANAAC AGNNGGCTGG CAACTAGAAG    61 GCACAGTCGA GGCTGATCAG CGGGTTTAAA CGGGCCCTCT AGACTCGAGC GGCCGCAAAC   121 GTTTTTTTTT TTTTTTTTTA TTTTAGGTGG CAGTTAGCCG TTACAAGAAG TTTTATTAAC   181 ATATTCTCTA ATAATCACAA ACCTATACAG CTATGTTCAG TATTTTCAAG GAGAGTGACT   241 TCTTACAAAA CGTTTATAGC CAATCATGGG ATTTTGCTAG AAAACGATGG CATATGCAAT   301 GACAGATTTG ACAGTTTAAT CAGAAAAATG GAATGATAGA AATGAAGCAG ACTCCCCACC   361 TACTGTGCTA ACTGGGTATG CAACAGTTTA ATGAAATACC ATAAGCCTTA GTTGGAGACT   421 GTGATGGAAC AGCGTANAGG CCATCCGTTT CCAGGCCATT CTGAGATCCA AGATGCCAAC   481 TCCTTGTTTC TATGAAACTT CACGAGATGT GCCAGTCAGA AATGGCATGC TGAACATTCT   541 GCAGAGCTGC AGCACTGGCC TNNNNNNNCA GGGCTACCAC ACTGGTGACA GTGCTGCTGG   601 TACCTGTGAT GGNTCTTCTG GTATCCATAT CTTATACNCA ACACCTATCC GTANNAAGAA   661 CTTCCGCANA ACTGCTCNNA NCTCNNGNAT CNNNTCNAAC TGCANNNTTT CNCACANGTN   721 NGGGNANNAN NTTGNANCNT GNGCTTTGNA CTTTTCNTCA TTTNNCNTNN NNNTTTANNT   781 NNNANTNNCN NCNNNANANN NGTNCNNNNN TNCNNANGNN NNTCNNANNN NCNNNGNTNN 841 ANTNNNNNNN NNN //
Primer: hBIG2_F1 (Pr0344) Region: insert mid (E738K) Sequence file: RDB15365_A7I5c.seq
>D03588D2_A7I5_2_hBIG2_F1_C09_09_ABI24.ab1     1 NNNNNNNNNN NNNNNNNTNC TCNNGGNNCA NGGCATGCTG GGAACGTCAG TTGAAGANAT    61 AGCCCAATTC CTGCACCAGG AGGAGCGCCT GGATTCCACC CAAGTAGGCG ATTTTCTGGG   121 AGATAGCGCA AGGTTCAACA AGGAGGTGAT GTATGCCTAC GTGGACCAAC TTGACTTCTG   181 TGAAAAAGAA TTTGTCTCAG CCCTGCGGAC ATTCCTAGAA GGTTTCCGCC TACCTGGGAA   241 AGCCCAGAAG ATCGATCGAT TAATGGAGAA GTTTGCCGCA AGATACATAG AATGCAACCA   301 AGGGCAAACT CTGTTTGCTA GTGCTGACAC TGCTTATGTC CTAGCGTATT CAATTATTAT   361 GCTGACTACA GACTTGCACA GTCCTCAGGT AAAAAATAAA ATGACGAAAG AGCAGTATAT   421 TAAAATGAAT CGGGGTATCA ATGATAGTAA AGATCTGCCA GAAGAGTATC TCTCAAGCAT   481 CTATGAAGAG ATAGAAGGCA AGAAAATTGC AATGAAAGAA ACAAAAGAGC TAACGATTGC   541 AACCAAATCT ACTAAGCAGA ATGTAGCTAG TGAAAAGCAG CGGCGGCTGC TGTACAACTT   601 AGAGATGGAG CAAATGGCTA AAACAGCCAA AGCTCTGATG GAGGCTGTGA GCCATGCCAA   661 AGCCCCGTTT ACCAGTGCCA CTCACCTGGA CCATGTCCGG CCAATGTTCA AACTGGTGTG   721 GACGCCACTA TTGGCAGCCT ACAGCATCGG ANTCCAGAAC TGTGANGANA CTGAANTGGC   781 CTCCTTGTGT TTGGAANGNA TCCGATGTGC AATCCGAATC GCNTGCATCT TTGGAATGCA   841 GCTNGAACGA NNNGNCNATG TTCNNNNNCT NGNTCGNTTC NNCNNANNNN CANCNNGNNN 901 CNGCNTCNCA NAAATGNANN CNNAAAAANN TCNNNNNNNN NN //
Primer: hBIG2_R1 (Pr1019) Region: insert 5', HA, Xpress tag, T7 tag, 6xHis, QBI SP163 Sequence file: RDB15365_A7I5d.seq
>D03588D2_A7I5_2_hBIG2_R1_H02_08_ABI08.ab1     1 NNNNNNNNNN NNNNNNNNNN AGCCTTGGCA GTGGTTTGAT TGATGAGNNT TTTGCTGGCC    61 AAATAGATAT TGTAACATGT TCTCACTGTC TGCAGGATAG TACCCTCATG AATTTCAATG   121 TGTGGGGAAG TCACTGCAGT CAGAAGAGCC TTAATTATTT GTAACTGAAC CCCTTCATCA   181 GTCTGAGGGC CCTGAAAACA ACTGCAAATG GTTTCAACAA TTCTGTCGAT CAGCCGCTTC   241 CCAGGGGCTC CACTGTCAGG GGCGTTGCCA GTGATGTGCC CGTATGCGAT GAGTTTCTGC   301 AAGCAGTCAA GGGATGTGCT GACTACCCTT GGGGACTTGG ACTGGCAAGC TAGCTCGAAT   361 GGAAGAAAAT ACTTGTCAGC TTCAATGAAG TTTGCCTTTG GTGGTGCAGC AGTGCCAAGC   421 CTCTGCTTTT CTATTTCTGC TTTAATTTCA TCGAGCGCCA CCTGGCAGGC CCTGCGCAGC   481 TGGGAGTGCT GGGGCCGCTT CACCTCCTTG TCGGCTAGGA TCTTCTCCAG GGCCCGGGAC   541 ACGAACATGC TCTTGGTCTG GCTCTCCTGC ATGGTGGATC CCGCGTAGTC GGGCACGTCG   601 TACGGGTACA TGGTAGGTAC CTTATCGTCA TCGTCGTACA GATCCCGACC CATTTGCTGT   661 CCACCAGTCA TGCTAGCCAT ACCATGATGA TGATGATGAT GAGAACCCCC CATGGTTTCG   721 GANGCCGTCC GGGGCCGGCG CGGCTCGCGC TCCCTCTCCG GCTCGGANTG CGAGGCAGCC   781 CGCTCTCTTG CGCGCCTTGG CGGGCTCCTC TCCCTTCTGG AACCGANGCC CGGGCCGGGG   841 CCTGNCTGCC GCTCGGCTGC CCCAAGCCTC TGCGCTNANC TTAAGTTTAA ACGCTAGNNA   901 GCTTGGGNCT CCCTATAGTG ANNNNTANTN NNTTCNANNA NCNNGTNNNC ANNGGGNNCT 961 CTANNNAGCN NNANANCTCT GCNNNNNNNN NN //
Primer: hBIG2_F2 (Pr1020) Region: insert 3' Sequence file: RDB15365_A7I5e.seq
>D03588D2_A7I5_2_hBIG2_F2_F02_06_ABI08.ab1     1 NNNNNNNNNN NGNNNNNNNN NNNNNTTCNN NAANNAGAAA CCNGCAGCCT GGCCTGTTGT    61 TTGAGGATCC TGTTTCGAAT GTATGTTGAT GAGAACCGCA GGGATTCCTG GGAAGAAATA   121 CAGCAGAGAC TTTTAACTGT TTGCAGTGAA GCTCTTGCCT ATTTCATCAC TGTGAATTCT   181 GAGAGCCATC GGGAGGCCTG GACAAGTCTC TTGTTGTTAC TTCTAACTAA AACCCTTAAG   241 ATAAATGATG AAAAGTTCAA AGCACATGCT TCAATGTACT ACCCCTACTT GTGTGAAATT   301 ATGCAGTTTG ACCTGATCCC TGAGCTCCGA GCAGTTCTGC GGAAGTTCTT CCTACGGATA   361 GGTGTTGTGT ATAAGATATG GATACCAGAA GAGCCATCAC AGGTACCAGC AGCACTGTCA   421 CCAGTGTGGT AGCCCTGGCT GCCCAGGCCA GTGCTGCAGC TCTGCAGAAT GTTCAGCATG   481 CCATTTCTGA CTGGCACATC TCGTGAAGTT TCATAGAAAC AAGGAGTTGG CATCTTGGAT   541 CTCAGAATGG CCTGGAAACG GATGGCCTCT ACGCTGTTCC ATCACAGTCT CCAACTAAGG   601 CTTATGGTAT TTCATTAAAC TGTTGCATAC CCAGTTAGCA CAGTAGGTGG GGAGTCTGCT   661 TCATTTCTAT CATTCCATTT TTCTGATTAA ACTGTCAAAT CTGTCATTGC ATATGCCATC   721 GTTTTCTAGC AAAATCCCAT GATTGNCTAT AAACGTTTTG TAAGAAGTCA CTCTCCTTGA   781 AAATACTGAA CATAGCTGTA TAGGTTTGTG ATTATTAGAG AATATGTTAA TAAAACTTCT   841 TGTAACGGCT AACTGCCACC TAAAATAAAA AAAAAAAAAA AAAACGTTNG CGGCNGCTCN   901 ANNCTANNNG NNCCNNTTAA ACCCGCTGAN CANNCNNNNN NNNGNCNNTN CNANTTNGNN 961 ANNNNTNNNN NNNNNN //
Primer: SV40pro_F (Pr0110) Region: EM7 pro, ZeoR Sequence file: RDB15365_A7I5f.seq
>D03588D2_A7I5_2_SV40pro_F_D09_12_ABI24.ab1     1 NNNNNNNNNN NNNNNNNNAN NNNTGNNGNN GCTTTTTTGG ANGCCTAGGC TTTTGCAAAA    61 AGCTCCCGGG AGCTTGTATA TCCNTTTTCG GATCTGATCA GCACGTGTTG ACAATTAATC   121 ATCGGCATAG TATATCGGCA TAGTATAATA CGACAAGGTG AGGAACTAAA CCATGGCCAA   181 GTTGACCAGT GCCGTTCCGG TGCTCACCGC GCGCGACGTC GCCGGAGCGG TCGAGTTCTG   241 GACCGACCGG CTCGGGTTCT CCCGGGACTT CGTGGAGGAC GACTTCGCCG GTGTGGTCCG   301 GGACGACGTG ACCCTGTTCA TCAGCGCGGT CCAGGACCAG GTGGTGCCGG ACAACACCCT   361 GGCCTGGGTG TGGGTGCGCG GCCTGGACGA GCTGTACGCC GAGTGGTCGG AGGTCGTGTC   421 CACGAACTTC CGGGACGCCT CCGGGCCGGC CATGACCGAG ATCGGCGAGC AGCCGTGGGG   481 GCGGGAGTTC GCCCTGCGCG ACCCGGCCGG CAACTGCGTG CACTTCGTGG CCGAGGAGCA   541 GGACTGACAC GTGCTACGAG ATTTCGATTC CACCGCCGCC TTCTATGAAA GGTTGGGCTT   601 CGGAATCGTT TTCCGGGACG CCGGCTGGAT GATCCTCCAG CGCGGGGNTC TCATGCTGGA   661 GTTCTTCGCC CACCCCAACT TGTTTATTGC AGCTTATAAT GNTTACNAAT AAAGCAATAG   721 CATCACAAAT TTCACAAATA AAGCATTTTT TTCACTGCAT TCTAGTTGTG GTTTGTCCNA   781 ACTCATCAAT GTATCTTATC ATGTCTGTAT ACCGNCNNNC TCTANCTANA NNTNNNNGNN   841 NNCATGGNCA TANNTGTTTC CNGNGNGAAN TNGNNNTCCG CNCACNNNTN CNNNNNNNNT 901 ACNANNCNNN NNNNATAAAG NGNNANNNCN NGGGNNNNNN NANNNNNNNN NN //

Please visit Sequencing and PCR primers for primer information.

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References

Original, user report and related articles

original	Ishizaki, R., Redundant roles of BIG2 and BIG1, guanine-nucleotide exchange factors for ADP-ribosylation factors in membrane traffic between the trans-Golgi network and endosomes. Mol. Biol. Cell 19 (6): 2650-2660 (2008). PMID 18417613. [link to RRC of NBRP]

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• human BIG2

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External Database
human BIG2

• in KEGG Gene
• in KEGG Orthology
• in NCBI Gene

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