PacBio sequencing of gene families - A case study with wheat gluten genes

TY - JOUR

T1 - PacBio sequencing of gene families - A case study with wheat gluten genes

AU - Zhang, Wei

AU - Ciclitira, Paul

AU - Messing, Joachim

PY - 2014/1/10

Y1 - 2014/1/10

N2 - Amino acids in wheat (Triticum aestivum) seeds mainly accumulate in storage proteins called gliadins and glutenins. Gliadins contain alpha/beta-, gamma- and omega-types whereas glutenins contain HMW- and LMW-types. Known gliadin and glutenin sequences were largely determined through cloning and sequencing by capillary electrophoresis. This time-consuming process prevents us to intensively study the variation of each orthologous gene copy among cultivars. The throughput and sequencing length of Pacific Bioscience RS (PacBio) single molecule sequencing platform make it feasible to construct contiguous and non-chimeric RNA sequences. We assembled 424 wheat storage protein transcripts from ten wheat cultivars by using just one single-molecule-real-time cell. The protein genes from wheat cultivar Chinese Spring are comparable to known sequences from NCBI. We demonstrated real-time sequencing of gene families with high-throughput and low-cost. This method can be applied to studies of gene amplification and copy number variation among species and cultivars. 

AB - Amino acids in wheat (Triticum aestivum) seeds mainly accumulate in storage proteins called gliadins and glutenins. Gliadins contain alpha/beta-, gamma- and omega-types whereas glutenins contain HMW- and LMW-types. Known gliadin and glutenin sequences were largely determined through cloning and sequencing by capillary electrophoresis. This time-consuming process prevents us to intensively study the variation of each orthologous gene copy among cultivars. The throughput and sequencing length of Pacific Bioscience RS (PacBio) single molecule sequencing platform make it feasible to construct contiguous and non-chimeric RNA sequences. We assembled 424 wheat storage protein transcripts from ten wheat cultivars by using just one single-molecule-real-time cell. The protein genes from wheat cultivar Chinese Spring are comparable to known sequences from NCBI. We demonstrated real-time sequencing of gene families with high-throughput and low-cost. This method can be applied to studies of gene amplification and copy number variation among species and cultivars. 

KW - PacBio sequencing

KW - Gene family

KW - Gluten diversity

KW - Copy number variation

KW - Repetitive sequence

KW - MAIZE

KW - ZEIN

KW - DNA

U2 - 10.1016/j.gene.2013.10.009

DO - 10.1016/j.gene.2013.10.009

M3 - Article

SN - 0378-1119

VL - 533

SP - 541

EP - 546

JO - Gene

JF - Gene

IS - 2

M1 - N/A

ER -