Name: NIA Mouse Embryonic Germ Cell cDNA Library (Long) Library ID: 1988 Organism: Mus musculus Strain: C57BL/6J Gender: male Age: 0 Stage: embryo Organ: germ cell Host: DH10B Vector: pCMV-SPORT6 Vector type: phagemid Insert digest: 5' SalI/NotI 3' Stop Codon Status: without Description: This is a long-transcript enriched cDNA library. Total RNA was obtained from Dr. Mark G. Carter (NIH/NIA-IRP). EG cells were cultured at 37C, 5%CO2 in DMEM supplemented with 15% ES cell-qualified FBS, 0.1mM non-essential amino acids, 2mM glutamine, penicillin/streptomycin, 1mM sodium pyruvate, 0.1mM beta- mercaptoethanol, and 10e7 units of LIF per liter. Double-stranded cDNAs were synthesized with an oligo-dT primer (5'- pGACTAGTTCTAGATCGCGAGCGGCCGCCCTTTTTTTTTTTTTTT-3')from 2.5 ug of total RNA, treated with T4 DNA polymerase, and purified by ethanol precipitation. The cDNAs were ligated to Lone-linker LL-Sal4 (5'-AAGAGGTAATCC-3'), purified by phenol/chloroform, and separated from free linker by Centricon 100. The cDNAs were amplified by long-range high fidelity PCR using Ex Taq polymerase (Takara) with primer Sal4-S. The cDNAs were digested with SalI and Not1I enzymes and cloned into SalI/NotI sites of pSPORT1 plasmid vector. The average insert size is about 4 kb. The library was constructed by Yulan Piao and Minoru Ko (NIH, NIA). Reference: Genome Res (2001) 11:1553-1558 [PMID: 11544199].