|Library name:||Human ORFeome|
|Library accession number:||LMBP LIB32|
|Available clones:||ORFeome entry clones|
|Vector sequence file:||p9210.txt|
|Vector circular map:||p9210.pdf|
|Vector accession number:||LMBP 9210|
|Vector type:||Gateway donor vector|
|Cloning sites:||Gateway BP reaction|
|Average insert size:||1126 bp|
|Distributed as:||individual clones as active cultures or plasmid DNA|
|Further information:||The host strain of the individual clones is Escherichia coli K12 DH5α
The individual clones can be grown in selective medium (LB-Lennox + 50 µg/ml spectinomycin) and cultivated at 37°C.
pENTR223-ORF (LMBP 9210) can be considered as a representative for all ORF plasmids available at BCCM/GeneCorner.
Related data can be found on: http://horfdb.dfci.harvard.edu/index.php?page=orfsearch
Sequences of the different ORFs can be found on: http://horfdb.dfci.harvard.edu/index.php?page=home
The library was based on the Mammalian Gene Collection (MGC), isolating all available ORFs in this resource via PCR, adding the Gateway attenuator sites attB1.1 and attB2.1 to the forward and reverse primer respectively.
The integrase binding sites within the attenuator sites attB1.1 and attB2.1 has been altered to more closely resemble the bacteriophage lambda consensus sequence, increasing the efficiency of Gateway recombination.
The forward ORF-specific primer includes the ATG start codon, and incorporates a Kozak consensus nucleotide G at position -3 relative to the ATG, thereby allowing efficient translation initiation from the ORF start codon.
The reverse ORF-specific primer does not contain the last nucleotide of the termination codon, to allow generation of C-terminal fusion proteins.
The resulting ORFs were cloned into pDONR223 (Invitrogen) via Gateway BP recombination.
Please take into account that the reading frame has been shifted by one nucleotide for the creation of C-terminal fusion proteins due to the absense of the last nucleotide of the ORF termination codon.
|History of deposit:||This plasmid was deposited by Dr M. Vidal1 2 3 and Dr D. Hill1 2 3. It was received from Dr I. Lemmens4.
1 Center for Cancer Systems Biology (CCSB), Boston, Massachusetts, USA
2 Department of Cancer Biology, Dana-Farber Cancer Institute, Boston, Massachusetts, USA
3 Department of Genetics, Harvard Medical School, Boston, Massachusetts, USA
4 Department of Medical Protein Research, VIB, Ghent, Belgium
|Library reference:||Yang et al., Nat. Methods 8 (2011), 659-661 [PMID: 21706014]|
|Related reference:||Rual et al., Genome Res. 14 (2004), 2128-2135 [PMID: 15489335]|
|Restricted distribution:||- BCCM MTA adapted for hORFeome clones
- The depositor will be informed of the customer's identity upon release of a sample outside the depositor's department or outside the departments in which BCCM/GeneCorner is embedded, namely UGent-DBMB and VIB-IRC.
- RECIPIENT agrees to refer to 'Yang et al., Nat. Methods 8 (2011), 659-661 [PMID: 21706014]' in the first publication making use of the MATERIAL.