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作物学报 ›› 2010, Vol. 36 ›› Issue (3): 435-441.doi: 10.3724/SP.J.1006.2010.00435

• 作物遗传育种·种质资源·分子遗传学 • 上一篇    下一篇

六个甘蓝型油菜油酸脱氢酶（FAD2）假基因的克隆和分析

肖钢1,2，张振乾1，邬贤梦1，谭太龙1,2，官春云1,*   

  1. 1湖南农业大学油料作物研究所/国家油料作物改良中心湖南分中心，2作物种质创新与资源利用国家重点实验室培育基地，湖南长沙410128
  • 收稿日期:2009-06-30 修回日期:2009-12-08 出版日期:2010-03-12 网络出版日期:2010-01-22
  • 通讯作者: 官春云,E-mail:guancy2000@yahoo.com.cn
  • 作者简介:肖钢，E-mail: sanjian123@yeah.net; Tel: 0731-4617941
  • 基金资助:

    本研究由国家重点基础研究发展计划（973计划）项目（2006CB101600）资助。

Cloning and Characterization of Six Oleic Acid Desaturase Pseudogenes of Brassica napus

XIAO Gang1,2,ZHANG Zhen-Qian1,WU Xian-Meng1,TAN Tai-Long1,2,GUAN Chun-Yun1*   

  1. 1 Oil Crops Institute/National Oil Crops Improvement Center, Hunau Agricultural University; 2 Pre-State Key Laboratory for Germplasm Innovation and Resource Utilization of Crops, Changsha 410128, China
  • Received:2009-06-30 Revised:2009-12-08 Published:2010-03-12 Published online:2010-01-22
  • Contact: GUAN Chun-Yun,E-mail:guancy2000@yahoo.com.cn
  • About author:XIAO Gang,E-mail: sanjian123@yeah.net; Tel: 0731-4617941

PDF

1939

被引次数

18

摘要：

以甘蓝型油菜湘油15为材料，采用PCR方法克隆并分析了56个FAD2基因克隆和47个FAD2基因cDNA克隆，从中发现6个新的FAD2基因拷贝。它们与公布的甘蓝型油菜FAD2基因(AY577313)具有87%以上的同源性，没有内含子，在开放阅读框中存在1~12个终止密码子，其中有2个拷贝具有转录功能。将这6个FAD2基因拷贝在酿酒酵母中进行体内表达实验，通过气相色谱检测脂肪酸组成证明其不具备油酸脱氢酶功能，与对照相比，也没有改变酵母体内脂肪酸组成。由此推测这6个FAD2基因拷贝为假基因。

关键词: 甘蓝型油菜, 油酸脱氢酶, FAD2, 基因表达, 酿酒酵母

Abstract:

The phenomenon of multi-copy genes is common in plants. Pseudogene is defined as an inactive gene, which can not synthesize functional proteins but share the similar DNA sequences with normal functional genes. In this study, 56 FAD2 DNA clones and 47 FAD2 seed cDNA clones of Brassica napus cv. Xiangyou 15 were investigated, and 6 new copies of FAD2 were detected, designated as FAD2P1-6 respectively. This sequence length of 6 copies ranged from 1 141–1 157 bp and there were no introns in their open reading frames (ORF). These 6 copies share 96.1% identity in nucleotides from one another, and share more than 87% nucleotides identity with AY577313. Deduced amino acid sequences revealed that 1–12 stop codons occurred in the coding region of six copies which will prevent them from coding for a functional protein. These six copies were investigated in vivo in Saccharomyces cerevisiae through being cloned into yeast expression vector pYES2.0, and the 16:2 and 18:2 fatty acids were determined by gas chromatographic analysis. The results revealed that the products of the six copies were not able to synthesize 16:2 and 18:2 fatty acids, suggesting that they are pseudogenes of FAD2. These multiple pseudogenes of FAD2 within the B. napus genome might result from the duplication of large chromosomal segments simultaneously following mutation. Because of the existence of multiple pseudogenes for FAD2 in B. napus genome, we should be careful in genetic research to identify true and false, to avoid wrong conclusions.

Key words: Brassica napus, FAD2, Oleic acid desaturase, Gene expression, Saccharomyces cerevisiae

[1] Okuley J, Lightner J, Feldmann K, Yadav N, Lark E, Browse J. Arabidopsis FAD2 gene encodes the enzyme that is essential for polyunsaturated lipid synthesis. Plant Cell, 1996, 6: 147-158

[2] Ohlrogge J, Jaworski J G. Regulation of fatty acid synthesis.Plant Mol Biol, 1997, 48: 109-136
[3] Nagahara U. Genome-analysis in Brassica with special reference to the experiments formation of B. napus and peculiar mode of fertilization. Jpn J Bot, 1935, 7: 389-452
[4] Scheffler J A, Sharpe A G, Schmidt H, Sperling P, Parkin I A P, Lühs W, Lydiate D J, Heinz E. Desaturase multigene families of Brassica napus arose through genome duplication.Theor Appl Genet, 1997, 94: 583-591

[5] Lynch M, Conery J S. The evolutionary fate and consequences of duplicate genes. Science, 2000, 290: 1151-1155

[6] Hirotsune S J, Yoshida N, Chen A, Garrett L, Sugiyama F, Takahashi S, Yagami K, Wynshaw-Boris A, Yoshiki A. An expressed pseudogene regulates the messenger-RNA stability of its homologous coding gene. Nature, 2003, 423: 91-96

[7] Frank A C, Amiri H, Andersson S G. Genome deterioration: Loss of repeated sequences and accumulation of junk DNA. Genetica, 2002 115(1): 1-12

[8] Wang G-L(王关林), Fang H-J(方宏筠). Plant Gene Engineering, 2nd edn (植物基因工程·第二版). Beijing: China Science Press, 2002. pp 744-745 (in Chinese)

[9] Wei D S, Li M C, Zhang X X, Ren Y, Xing L J. Identification and characterization of a novel Δ12-fatty acid desaturase gene from Rhizopus arrhizus. FEBS Lett, 2004, 573: 45-50
[10] Romanos M A, Scorer C A, Clare J J. Foreign gene expression in yeast: A review. YEAST, 1992, 8: 423-488

[11] Niu B, Ye H X, Xu Y, Wang S H, Chen P, Peng S M, Ou Y C,Tang L,Chen F. Cloning and characterization of a novel Δ12-fatty acid desaturase gene from the tree Sapium sebiferum.Biotechnol Lett, 2007, 29: 959-964

[12] Guan C-Y(官春云). Advance of high oleic acid in oil seed breeding. Crop Res (作物研究), 2006, 20(1): 1-8 (in Chinese)

[13] Jacq C, Miller J R, Brownlee G G. A pseudogene structure in 5S DNA of Xenopus laevis. Cell, 1977, 13: 109-120

[14] Zhang Z L, Harrison P M, Liu Y, Gerstein M. Millions of years of evolution preserved: A comprehensive catalog of the processed pseudogenes in the human genome.Genome Res,2003,13: 2541-2558

[15] Lynch M, Conery J S. The evolutionary fate and consequences of duplicate genes. Science, 2000, 290: 1151-1155
[16] Balakirev E S, Ayala F J. Pseudogenes: Are they “junk” or “functional” DNA.
Annu Rev Genet, 2003, 37: 123-151
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