title: An efficient approach for constructing shRNA expression vectors based on short oligonucleotide synthesis.
abstract: Traditional strategies for establishing shRNA expression constructs are inefficient, error-prone, or costly. We describe a simple approach that overcomes these drawbacks. Briefly, the sense and antisense strands of the short hairpin RNA coding sequence are segmented into two parts, respectively, at asymmetric sites. The four resulting short oligonucleotides are synthesized. Each oligonucleotide is annealed with its opposite, resulting in a double-stranded fragment with sticky termini at both ends. The two fragments so generated can be easily spliced by simple ligation to reconstitute the full-length short hairpin RNA coding sequence which can then be cloned into an appropriately restricted vector.
authors.affiliation: Key Lab of Ministry of Health for Hormone and Development, Institute of Endocrinology, Tianjin Medical University, Tianjin 300070, People's Republic of China.
authors.forename: Xiao-Xia
authors.initials: XX
authors.lastname: Li
authors.forename: Hong-Wei
authors.initials: HW
authors.lastname: Jia
authors.forename: Jin-Xing
authors.initials: JX
authors.lastname: Quan
authors.forename: Yan-Li
authors.initials: YL
authors.lastname: Jiao
authors.forename: Yu-Hong
authors.initials: YH
authors.lastname: Yang
authors.forename: Bao-Li
authors.initials: BL
authors.lastname: Wang
authors.forename: Zhi
authors.initials: Z
authors.lastname: Yao
mesh_terms: D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D005786:Gene Expression Regulation; D005822:Genetic Vectors / Q000235:genetics*; D037181:Lectins, C-Type / Q000235:genetics / Q000378:metabolism; D008967:Molecular Biology / Q000379:methods*; D008969:Molecular Sequence Data; D009841:Oligonucleotides / Q000096:biosynthesis*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D034741:RNA, Small Interfering / Q000235:genetics*; D051381:Rats; D015183:Restriction Mapping
chemical_list: D037181:Lectins, C-Type; D009841:Oligonucleotides; D012333:RNA, Messenger; D034741:RNA, Small Interfering