Construction Of CDNA Library Of Mouse Olfactory Bulb

The olfactory system is an unusual tissue in that it can support neurogenesis throughoutlifetime, permitting the in-growth and synapse formation of olfactory recptor axons into thecentral nervous system (CNS) environment of the olfactory bulb (OB). It is thought that thisunusual property is in due to the olfactory glial cells, termed olfactory ensheathing cells(OECs). OECs originate from the placode and possess many properties in commom with glialcells from the peripheral nervous system (PNS), Schwann cells. The presence of olfactoryensheathing cells in the OB might account for regeneration of injured axons. Olfactoryensheathing cells enfold regenerating olfactory axons along the entire path, preventing themfrom contacting any other glial cell type, and most likely creating the appropriateenvironment for their growth.Recent data has suggested that olfactory ensheathing cells are a distinct glial cell typeand possess properties, which might make them more suitable for transplant-mediated repairof central nervous system injury models. It was not clear how OECs promote CNSregeneration so we constructed a cDNA library using olfactory bulb of mouse for in serch ofspecific gene expessed in OECs.Total RNA were isolated from OB of mouse using Nucleospin RNA Ⅱkit. First-strandcDNAs were synthesized using SMART Ⅳ Oligonucleotide and CDS Ⅲ/3′PCR Primer asprimers. Double-stranded cDNAs were synthesized using LD-PCR method. The dscDNAswere digested by Proteinase K and Sfi Ⅰenzyme, less than 500bp fragments were separatedby CHROMA SPIN-400 column to ensure the length of inserts, then the cDNAs were ligatedto the Sfi digested λTripl Ex2 vector. After packaging, the OB cDNA library was constructed.The titer of the primary library was 1.3×106pfu/ml, the recombination was 93%, the size ofthe inserts were between 200bp-3kb, the titer of the amplified OB cDNA library was1×109pfu/ml. These results indicate that the quality of this cDNA library is good enough forfurther cloning of the OECs specific expression genes.