title: A-T linker adapter polymerase chain reaction for determining flanking sequences by rescuing inverse PCR or thermal asymmetric interlaced PCR products.
abstract: The polymerase chain reaction (PCR)-based genome walking method has been extensively used to isolate unknown flanking sequences, whereas nonspecific products are always inevitable. To resolve these problems, we developed a new strategy to isolate the unknown flanking sequences by combining A-T linker adapter PCR with inverse PCR (I-PCR) or thermal asymmetric interlaced PCR (TAIL-PCR). The result showed that this method can be efficiently achieved with the flanking sequence from the Arabidopsis mutant and papain gene. Our study provides researchers with an additional method for determining genomic DNA flanking sequences to identify the target band from bulk of bands and to eliminate the cloning step for sequencing.
authors.affiliation: Laboratory of Food Safety, College of Food Science and Nutritional Engineering, China Agricultural University, Beijing 100083, China.
authors.forename: Quoclinh
authors.initials: Q
authors.lastname: Trinh
authors.affiliation: Laboratory of Food Safety, College of Food Science and Nutritional Engineering, China Agricultural University, Beijing 100083, China.
authors.forename: Pengyu
authors.initials: P
authors.lastname: Zhu
authors.affiliation: Laboratory of Food Safety, College of Food Science and Nutritional Engineering, China Agricultural University, Beijing 100083, China.
authors.forename: Hui
authors.initials: H
authors.lastname: Shi
authors.affiliation: Laboratory of Food Safety, College of Food Science and Nutritional Engineering, China Agricultural University, Beijing 100083, China. Electronic address: xuwentaoboy@sina.com.
authors.forename: Wentao
authors.initials: W
authors.lastname: Xu
authors.affiliation: Laboratory of Food Safety, College of Food Science and Nutritional Engineering, China Agricultural University, Beijing 100083, China.
authors.forename: Junran
authors.initials: J
authors.lastname: Hao
authors.affiliation: Laboratory of Food Safety, College of Food Science and Nutritional Engineering, China Agricultural University, Beijing 100083, China; Supervision, Inspection, and Testing Center of Genetically Modified Organisms, Ministry of Agriculture, Beijing 100083, China.
authors.forename: Yunbo
authors.initials: Y
authors.lastname: Luo
authors.affiliation: Laboratory of Food Safety, College of Food Science and Nutritional Engineering, China Agricultural University, Beijing 100083, China; Supervision, Inspection, and Testing Center of Genetically Modified Organisms, Ministry of Agriculture, Beijing 100083, China.
authors.forename: Kunlun
authors.initials: K
authors.lastname: Huang
mesh_terms: D024506:5' Flanking Region / Q000235:genetics*; D016386:Chromosome Walking; D010206:Papain / Q000235:genetics; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic / Q000235:genetics; D017422:Sequence Analysis, DNA / Q000379:methods*
chemical_list: D010206:Papain
keywords: A-T linker adapter PCR; Flanking sequence; Genome walking; PCR product rescue