Ptcorp gene induced by cold stress was identified by proteomic analysis in leaves of Poncirus trifoliata (L.) Raf.
A proteomic approach was employed to investigate the cold stress-responsive proteins in trifoliate orange (Poncirus trifoliata (L.) Raf.), which is a well-known cold tolerant citrus relative and widely used as rootstock in China. Two-year-old potted seedlings were exposed to freezing temperature (-6°C) for 50 min (nonlethal) and 80 min (lethal), and the total proteins were isolated from leaves of the treated plants. Nine differentially accumulated proteins over 2-fold changes in abundance were identified by two-dimensional gel electrophoresis and mass spectrometry. Among these proteins, a resistance protein induced by the nonlethal cold treatment (protein spot #2 from P. trifoliata) was selected as target sequence for degenerated primer design. By using the designed primers, a PCR product of about 700 bp size was amplified from P. trifoliata genomic DNA, which was further cloned and sequenced. A nucleotide sequence of 676 bp was obtained and named Ptcorp. Blast retrieval showed that Ptcorp shared 88% homology with an EST of cold acclimated Bluecrop (Vaccinium corymbosum) library (Accession number: CF811080), indicating that Ptcorp had association with cold acclimation. Semiquantitative RT-PCR analysis demonstrated that Ptcorp gene was up-regulated by cold stress which was consistent with the former result of protein expression profile. As the resistance protein (NBS-LRR disease resistance protein family) gene was up-regulated by cold stress in trifoliate orange and satsuma mandarin, it may imply that NBS-LRR genes might be associated with cold resistance in citrus.
College of Bioscience and Biotechnology, Hunan Agricultural University, Changsha, 410128, Hunan, China. longgy63@163.com
Guiyou
G
Long
Jinyu
J
Song
Ziniu
Z
Deng
Jie
J
Liu
Liqun
L
Rao
D000595:Amino Acid Sequence; D003080:Cold Temperature; D017931:DNA Primers / Q000378:metabolism; D018744:DNA, Plant / Q000235:genetics; D060467:Disease Resistance / Q000235:genetics; D015180:Electrophoresis, Gel, Two-Dimensional; D020869:Gene Expression Profiling; D018506:Gene Expression Regulation, Plant; D017343:Genes, Plant / Q000235:genetics*; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D010935:Plant Diseases / Q000235:genetics; D018515:Plant Leaves / Q000235:genetics*; D010940:Plant Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D032104:Poncirus / Q000235:genetics* / Q000502:physiology; D040901:Proteomics / Q000379:methods*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012984:Software; D013312:Stress, Physiological / Q000235:genetics*
D017931:DNA Primers; D018744:DNA, Plant; D010940:Plant Proteins