International Crops Research Institute for the Semi-Arid Tropics (ICRISAT), Patancheru, Greater Hyderabad, AP, India.
BMC Genomics. 2009 Nov 15;10:523. doi: 10.1186/1471-2164-10-523.
Chickpea (Cicer arietinum L.), an important grain legume crop of the world is seriously challenged by terminal drought and salinity stresses. However, very limited number of molecular markers and candidate genes are available for undertaking molecular breeding in chickpea to tackle these stresses. This study reports generation and analysis of comprehensive resource of drought- and salinity-responsive expressed sequence tags (ESTs) and gene-based markers.
A total of 20,162 (18,435 high quality) drought- and salinity- responsive ESTs were generated from ten different root tissue cDNA libraries of chickpea. Sequence editing, clustering and assembly analysis resulted in 6,404 unigenes (1,590 contigs and 4,814 singletons). Functional annotation of unigenes based on BLASTX analysis showed that 46.3% (2,965) had significant similarity (< or =1E-05) to sequences in the non-redundant UniProt database. BLASTN analysis of unique sequences with ESTs of four legume species (Medicago, Lotus, soybean and groundnut) and three model plant species (rice, Arabidopsis and poplar) provided insights on conserved genes across legumes as well as novel transcripts for chickpea. Of 2,965 (46.3%) significant unigenes, only 2,071 (32.3%) unigenes could be functionally categorised according to Gene Ontology (GO) descriptions. A total of 2,029 sequences containing 3,728 simple sequence repeats (SSRs) were identified and 177 new EST-SSR markers were developed. Experimental validation of a set of 77 SSR markers on 24 genotypes revealed 230 alleles with an average of 4.6 alleles per marker and average polymorphism information content (PIC) value of 0.43. Besides SSR markers, 21,405 high confidence single nucleotide polymorphisms (SNPs) in 742 contigs (with > or = 5 ESTs) were also identified. Recognition sites for restriction enzymes were identified for 7,884 SNPs in 240 contigs. Hierarchical clustering of 105 selected contigs provided clues about stress- responsive candidate genes and their expression profile showed predominance in specific stress-challenged libraries.
Generated set of chickpea ESTs serves as a resource of high quality transcripts for gene discovery and development of functional markers associated with abiotic stress tolerance that will be helpful to facilitate chickpea breeding. Mapping of gene-based markers in chickpea will also add more anchoring points to align genomes of chickpea and other legume species.
鹰嘴豆（Cicer arietinum L.）是世界上重要的粮食豆类作物，受到干旱和盐胁迫的严重威胁。然而，在鹰嘴豆中开展分子育种以应对这些胁迫的分子标记和候选基因非常有限。本研究报道了干旱和盐胁迫响应表达序列标签（ESTs）和基于基因标记的综合资源的产生和分析。
从鹰嘴豆的十个不同根组织 cDNA 文库中总共生成了 20162 个（18435 个高质量）干旱和盐胁迫响应 ESTs。通过序列编辑、聚类和组装分析，得到了 6404 个基因（1590 个串联和 4814 个单核苷酸）。基于 BLASTX 分析的基因功能注释显示，46.3%（2965 个）与非冗余 UniProt 数据库中的序列具有显著相似性（<或=1E-05）。与四种豆科植物（Medicago、Lotus、大豆和落花生）和三种模式植物（水稻、拟南芥和杨树）的 ESTs 进行 BLASTN 分析，为豆科植物之间的保守基因以及鹰嘴豆的新转录本提供了见解。在 2965 个（46.3%）显著基因中，只有 2071 个（32.3%）基因可以根据基因本体论（GO）描述进行功能分类。共鉴定到 2029 个含有 3728 个简单重复序列（SSR）的序列，开发了 177 个新的 EST-SSR 标记。在 24 个基因型上对 77 个 SSR 标记的一组实验验证显示，平均每个标记有 4.6 个等位基因，平均多态信息含量（PIC）值为 0.43。除了 SSR 标记外，还在 742 个包含>或=5 个 EST 的 contigs 中鉴定到 21405 个高置信度单核苷酸多态性（SNP）。在 240 个 contigs 中的 7884 个 SNP 中识别到了限制酶识别位点。105 个选定 contigs 的层次聚类提供了关于胁迫响应候选基因的线索，其表达谱显示在特定胁迫文库中占主导地位。
生成的鹰嘴豆 EST 集可作为基因发现和与非生物胁迫耐受性相关的功能标记开发的高质量转录本资源，这将有助于促进鹰嘴豆的育种。在鹰嘴豆中对基于基因的标记进行作图，也将为鹰嘴豆和其他豆科植物的基因组对齐增加更多的锚定点。
