Establishment and Optimization of ISSR-PCR Reaction System of Machilus thunbergii

JIANG Rong-bo; JIANG Jing-min; LIU Jun; CHEN Yi-tai; LUAN Qi-fu; YUE Hua-feng

Volume 24 Issue 2

Article Contents

Turn off MathJax

Article Navigation > Forest Research > 2011 > 24(2): 194-199

Citation:

Establishment and Optimization of ISSR-PCR Reaction System of Machilus thunbergii

1.
Research Institute of Subtropical Forestry, Chinese Academy of Forestry, Fuyang 311400, Zhejiang, China
2.
Paulownia Research and Development Center, State Forestry Administration, Zhengzhou 450003, He’nan, China

Received Date: 2010-05-19

Abstract

In order to establish stable ISSR-PCR reaction system of Machilus thunbergii, single factor and orthogonal experiment design were used to optimize the reaction system after exploring M. thunbergii DNA extraction. For finding optimal concentration of factors of ISSR-PCR, the different levels of concentration of Mg2+, primer, template DNA, dNTPs were trailed by single factor experiment. Meanwhile, in order to improve the reliability of the results of the single-factor test, the authors also adopted orthogonal design by four factors, three levels for further optimizing and screening the best conditions of Mg2+, primer, template DNA, and dNTPs. The best ISSR-PCR reaction system of M. thunbergii was eventually established by comparing the two method’s results, which included 20 μL reaction system, Taq enzyme, 0.05 U·μL-1,Mg2+ 2.0 mmol·L-1 template DNA 1 ng·L-1,dNTPs 0.3 mmol·L-1,primers(835) 0.5 μmol·L-1, and 1×PCR buffer. The establishment of the better repeatability and stability ISSR-PCR reaction system could provide technical support for further research of genetic structure and genetic variety of M. thunbergii group.
- Machilus thunbergii,
- ISSR-PCR reaction polymorphism,
- single factor experiments,
- orthogonal design

References

[1]	谢力文.红楠无性繁殖技术[J]. 林业实用技术,2005(9): 45-46
[2]	江香梅,俞湘. 红楠及其研究进展[J]. 江西农业大学学报, 2001, 23(2): 231-235
[3]	Maesako Y. Community structure of Machilus thunbergii forests disturbed by birds (Calonectris leucomelas: streaked shearwater) on Kanmurijima island, Kyoto Prefecture, Japan[J]. Japanese Journal of Ecology, 1985,35(3): 387-400
[4]	Kim C S,Cho R M, Kim W W. Heritabilities for height and diameter at root collar and delermenation of proportion of selection for estimation of genetic gain in 4-yr-old open-pollinated progenies of Machilus thunbergii[J]. Research Report of the Institute of Forest Genetles,1992, 28: 40-47
[5]	邵春荣,周芳勇,魏斌,等.红楠播种育苗试验研究[J].林业科技开发, 2007,21(2): 73-76
[6]	曾繁茂.红楠群落特征的初步研究[J].福建林业科技,1999,26(1): 42-45
[7]	陈艳,沈浪,应向阳,等.中国大陆青冈分布区东缘大金山岛种群种子形态变异[J].广西植物, 2007, 27(4): 555-559
[8]	Bornet B, Branchard M. Nonanchored inter simple sequence repeat (ISSR) markers: reproducible and specific tools for genome finger-printing[J]. Plant Molecular Biology Reporter, 2001, 19: 209-215
[9]	Sankar A A, Moore G A. Evaluation of inter-simple sequence repeat analysis for mapp ing in Citrus and extension of the genetic linkage map[J]. Theor Appl Genet, 2001, 102(2-3): 206-214
[10]	Ratnaparkhe M B, Santra D K, Tullu A, et al. Inheritance of inter-simple-sequence-repeat polymorphisms and linkage with a fusariumwilt resistance gene in chickpea[J]. Theor Appl Genet, 1998, 96(3-4): 348-353
[11]	Luan S, Chiang T Y, Gong X. High genetic diversity vs. low genetic differentiation in Nouelia insignis(Asteraceae), a narrowly distributed and endemic species in China, revealed by ISSR fingerp rinting[J]. Annals of Botany, 2006, 98(3): 583-589
[12]	Escandón A S, Zelener N, de la Torre M P, et al. Molecular identification of new varieties of Nierembergia linariaefolia(Graham) , a native Argentinean ornamental plant[J]. J Appl Genet, 2007, 48(2) : 115-123
[13]	续九如,黄智慧. 林业试验设计 . 北京: 中国林业出版社, 1998 Doyle J J, Doyle J L. A rapid DNA isolation procedure for small quantities of fresh leaf tissue[J]. Phytochem Bull, 1987, 19: 11-15
[14]	刘军. 毛红椿天然居群遗传结构的研究 . 北京: 中国林业科学研究院,2007
[15]	高大伟. 樟科植物DNA Barcode及香樟系统地理学的初步研究 . 上海: 华东师范大学,2008
[16]	徐虹,郑敏,章军,等. 3种樟科植物的细胞总DNA提取[J]. 云南植物研究,2004, 26 (4) : 451-457
[17]	叶倩. 夏腊梅、红楠的种质特异性分子标记的研究 . 杭州: 浙江大学,2006
[18]	Reddym P, Saril N, Siddiq E A. Inter simple sequence repeat (ISSR) polymorphism and its application in plant breeding[J] . Euphytica, 2002,128 (1) : 9-17
[19]	谢运海,夏德安,姜静,等. 利用正交设计优化水曲柳ISSR-PCR反应体系[J]. 分子植物育种,2005,3(3): 445-450
[20]	李鹏, 汪阳东,陈益存,等. 油桐ISSR-PCR最佳反应体系的建立[J]. 林业科学研究,2008, 21(2) : 194-199
[21]	余艳, 陈海山, 葛学军,等. 简单重复序列区间(ISSR)引物反应条件优化与筛选[J]. 热带亚热带植物学报,2003,11 (1):15-19
[22]	乔玉山, 章镇, 房经贵,等. 李种质资源ISSR反应体系的建立[J] . 果树学报, 2003, 20 (4) : 270-274
[23]	杨华, 宋绪忠, 尹光天,等. 黄藤ISSR反应体系的条件优化[J]. 福建林学院学报,2006, 26 (2) : 152-155

Proportional views

通讯作者: 陈斌, bchen63@163.com

1.
沈阳化工大学材料科学与工程学院沈阳 110142

Get Citation

PDF

XML

Article views(3431) PDF downloads(1424) Cited by()

Proportional views

Establishment and Optimization of ISSR-PCR Reaction System of Machilus thunbergii

1. Research Institute of Subtropical Forestry, Chinese Academy of Forestry, Fuyang 311400, Zhejiang, China

2. Paulownia Research and Development Center, State Forestry Administration, Zhengzhou 450003, He’nan, China

Received Date: 2010-05-19

Abstract: In order to establish stable ISSR-PCR reaction system of Machilus thunbergii, single factor and orthogonal experiment design were used to optimize the reaction system after exploring M. thunbergii DNA extraction. For finding optimal concentration of factors of ISSR-PCR, the different levels of concentration of Mg2+, primer, template DNA, dNTPs were trailed by single factor experiment. Meanwhile, in order to improve the reliability of the results of the single-factor test, the authors also adopted orthogonal design by four factors, three levels for further optimizing and screening the best conditions of Mg2+, primer, template DNA, and dNTPs. The best ISSR-PCR reaction system of M. thunbergii was eventually established by comparing the two method’s results, which included 20 μL reaction system, Taq enzyme, 0.05 U·μL-1,Mg2+ 2.0 mmol·L-1 template DNA 1 ng·L-1,dNTPs 0.3 mmol·L-1,primers(835) 0.5 μmol·L-1, and 1×PCR buffer. The establishment of the better repeatability and stability ISSR-PCR reaction system could provide technical support for further research of genetic structure and genetic variety of M. thunbergii group.

HTML

Reference (23)

[1]	谢力文.红楠无性繁殖技术[J]. 林业实用技术,2005(9): 45-46
[2]	江香梅,俞湘. 红楠及其研究进展[J]. 江西农业大学学报, 2001, 23(2): 231-235
[3]	Maesako Y. Community structure of Machilus thunbergii forests disturbed by birds (Calonectris leucomelas: streaked shearwater) on Kanmurijima island, Kyoto Prefecture, Japan[J]. Japanese Journal of Ecology, 1985,35(3): 387-400
[4]	Kim C S,Cho R M, Kim W W. Heritabilities for height and diameter at root collar and delermenation of proportion of selection for estimation of genetic gain in 4-yr-old open-pollinated progenies of Machilus thunbergii[J]. Research Report of the Institute of Forest Genetles,1992, 28: 40-47
[5]	邵春荣,周芳勇,魏斌,等.红楠播种育苗试验研究[J].林业科技开发, 2007,21(2): 73-76
[6]	曾繁茂.红楠群落特征的初步研究[J].福建林业科技,1999,26(1): 42-45
[7]	陈艳,沈浪,应向阳,等.中国大陆青冈分布区东缘大金山岛种群种子形态变异[J].广西植物, 2007, 27(4): 555-559
[8]	Bornet B, Branchard M. Nonanchored inter simple sequence repeat (ISSR) markers: reproducible and specific tools for genome finger-printing[J]. Plant Molecular Biology Reporter, 2001, 19: 209-215
[9]	Sankar A A, Moore G A. Evaluation of inter-simple sequence repeat analysis for mapp ing in Citrus and extension of the genetic linkage map[J]. Theor Appl Genet, 2001, 102(2-3): 206-214
[10]	Ratnaparkhe M B, Santra D K, Tullu A, et al. Inheritance of inter-simple-sequence-repeat polymorphisms and linkage with a fusariumwilt resistance gene in chickpea[J]. Theor Appl Genet, 1998, 96(3-4): 348-353
[11]	Luan S, Chiang T Y, Gong X. High genetic diversity vs. low genetic differentiation in Nouelia insignis(Asteraceae), a narrowly distributed and endemic species in China, revealed by ISSR fingerp rinting[J]. Annals of Botany, 2006, 98(3): 583-589
[12]	Escandón A S, Zelener N, de la Torre M P, et al. Molecular identification of new varieties of Nierembergia linariaefolia(Graham) , a native Argentinean ornamental plant[J]. J Appl Genet, 2007, 48(2) : 115-123
[13]	续九如,黄智慧. 林业试验设计 . 北京: 中国林业出版社, 1998 Doyle J J, Doyle J L. A rapid DNA isolation procedure for small quantities of fresh leaf tissue[J]. Phytochem Bull, 1987, 19: 11-15
[14]	刘军. 毛红椿天然居群遗传结构的研究 . 北京: 中国林业科学研究院,2007
[15]	高大伟. 樟科植物DNA Barcode及香樟系统地理学的初步研究 . 上海: 华东师范大学,2008
[16]	徐虹,郑敏,章军,等. 3种樟科植物的细胞总DNA提取[J]. 云南植物研究,2004, 26 (4) : 451-457
[17]	叶倩. 夏腊梅、红楠的种质特异性分子标记的研究 . 杭州: 浙江大学,2006
[18]	Reddym P, Saril N, Siddiq E A. Inter simple sequence repeat (ISSR) polymorphism and its application in plant breeding[J] . Euphytica, 2002,128 (1) : 9-17
[19]	谢运海,夏德安,姜静,等. 利用正交设计优化水曲柳ISSR-PCR反应体系[J]. 分子植物育种,2005,3(3): 445-450
[20]	李鹏, 汪阳东,陈益存,等. 油桐ISSR-PCR最佳反应体系的建立[J]. 林业科学研究,2008, 21(2) : 194-199
[21]	余艳, 陈海山, 葛学军,等. 简单重复序列区间(ISSR)引物反应条件优化与筛选[J]. 热带亚热带植物学报,2003,11 (1):15-19
[22]	乔玉山, 章镇, 房经贵,等. 李种质资源ISSR反应体系的建立[J] . 果树学报, 2003, 20 (4) : 270-274
[23]	杨华, 宋绪忠, 尹光天,等. 黄藤ISSR反应体系的条件优化[J]. 福建林学院学报,2006, 26 (2) : 152-155

Establishment and Optimization of ISSR-PCR Reaction System of Machilus thunbergii

Abstract

References

Proportional views

通讯作者: 陈斌, bchen63@163.com

Proportional views

Establishment and Optimization of ISSR-PCR Reaction System of Machilus thunbergii

HTML

Catalog

Export File

Citation

Format

Content