Chen Yu-Ning, Wei Wen-Hui, Ren Xiao-Ping, Zhao Xue-Ya, Zhou Xiao-Jing, Huang Li, Tang Xing-Chun, Jiang Hui-Fang
Oil Crops Research Institute of the Chinese Academy of Agricultural Sciences/Key Laboratory of Biology and Genetic Improvement of Oil Crops, Ministry of Agriculture, Wuhan, 430062, China.
School of Life Sciences, Hubei University, Wuhan, 430062, China.
Bot Stud. 2014 Dec;55(1):8. doi: 10.1186/1999-3110-55-8. Epub 2014 Jan 19.
Arachis hypogaea L. (2n = 4× = 40, AABB) is one of the most important oil and economic crop plants in the word. This species has the largest genome size of about 2,813 Mb among the oil crop species. Zhonghua 8 is a peanut cultivar planted widely in central China and has several superior traits including high oil content, high yield and disease resistance. A high-quality BAC library of Zhonghua 8 was constructed for future researches on the genomics of Chinese peanut cultivars.
A Hin d III-digested genomic BAC (bacterial artificial chromosome) library was constructed with the genomic DNA from leaves of Zhonghua 8. This BAC library consists of 160,512 clones and the average insert is estimated about 102 kb ranging from 30 to 150 kb. The library represents about 5.55× haploid genome equivalents, and provides a 99.71% probability of finding specific genes. The empty-vector rate is under 5 percent detected from 200 randomly selected clones. Probing of 384 clones with the psbA gene of barley chloroplast and the atp6 gene of rice mitochondrion indicated that the contamination with organellar DNA is insignificant. Successive subculture of three clones showed that the inserts are stable in one hundred generations.
This study presented the construction of a high-quality BAC library for the genome of Chinese cultivated peanut. Many essential experiences were summarized in the present study. This BAC library can serve as a substantial platform for development of molecular marker, isolation of genes and further genome research.
花生(Arachis hypogaea L.,2n = 4× = 40,AABB)是世界上最重要的油料和经济作物之一。在油料作物中,该物种的基因组最大,约为2813 Mb。中花8号是一种在中国中部广泛种植的花生品种,具有含油量高、产量高和抗病等多个优良性状。为了未来对中国花生品种进行基因组学研究,构建了高质量的中花8号BAC文库。
利用中花8号叶片的基因组DNA构建了一个经Hin d III酶切的基因组BAC(细菌人工染色体)文库。该BAC文库由160,512个克隆组成,平均插入片段估计约为102 kb,范围在30至150 kb之间。该文库代表约5.55倍单倍体基因组当量,找到特定基因的概率为99.71%。从随机挑选的200个克隆中检测到的空载率低于5%。用大麦叶绿体的psbA基因和水稻线粒体的atp6基因对384个克隆进行探针杂交,结果表明细胞器DNA的污染不显著。对三个克隆进行连续传代培养表明,插入片段在100代中是稳定的。
本研究展示了中国栽培花生基因组高质量BAC文库的构建。本研究总结了许多重要经验。该BAC文库可作为开发分子标记、分离基因和进一步进行基因组研究的重要平台。