cDNA Library, S.pombe, After + HU, γ, MMS

cDNA Library, S.pombe, After + HU, γ, MMS

Cat. No. FNK-02-707
Store at -20°C
Size 500 ng
Quantity 

500 ng (40 ng/ul, 13ul) in 10 mM Tris-HCl-1mM EDTA (pH 7.5)

Quality

1. Number of independent clones: 7.7 x 10⁶
2. Average insert size : longer than 1 kb 

Description

This cDNA library (plasmid DNA) is constructed from poly(A)+ RNA derived from Schizosaccharomyces pombe, strain h-L972 after treatment with HU(hydroxyurea), γ-radiation, and MMS(methylmethane sulfonate). It is constructed by the Linker-Primer method (Ref.1) by Professor Hiroshi Nojima of the Research Institute for Microbial Diseases, Osaka University. This library is unidirectionally cloned using the oligo (dT)18 linker primer which contains the restriction enzyme site of Not I, and BamHI (Bgl II)-Sma I adaptor. The pLZ3 vector used in this library can replicate both in S. pombe and in E. coli, and express cloned genes not only in S. pombe but also in mammalian cells as it contains the SV40 promoter. It also contains f1 ori which is necessary for ssDNA synthesis, and bacteriophage T7 and T3 promoter for RNA synthesis (see Ref.2). 

Applications

1. PCR screening of known or unknown gene: Prepare the primers for the known or unknown gene (cDNA) and amplify the gene by PCR from this library followed by cloning to an appropriate vector. It is useful for large-scale protein productions, and preparation of probes, etc. Standard amplifying conditions: 35 cycles of PCR reactions using 10-100 ng of cDNA as a template. (Change the quantity of template and the number of cycles depending on the expression rate of mRNA of the objective gene.)
2. Cloning the cDNA by functional complementation of the corresponding mutant strains. 

Note

* This library is to be used only by the researches of the purchaser. It is not allowed to amplify and transfer the library to a third person.
* Related products: human tissue specific cDNA libraries and cDNA libraries of model organisms

References

1. Kobori M et al ” Large scale isolation of osteoclast-specific genes by an improved method involving the preparation of a subtracted cDNA library.” Genes Cells 3: 459-475 (1998) PMID: 9753427
2. Tanaka S and Nojima H ”Nik1: a Nim1-like protein kinase of S. cerevisiae interacts with the Cdc28 complex and regulates cell cycle progression.” Genes Cells 1, 905-921 (1996) PMID: 9077450
3. Sambrook J and Russell DW Molecular Cloning Chapter 11 ”Preparation of cDNA libraries and gene identification.” CSHL Press (2001)