大麻ISSR反应体系的优化与引物的初步筛选

doi:10.11924/j.issn.1000-6850.2013-2549

中国农学通报 ›› 2014, Vol. 30 ›› Issue (12): 105-109.doi: 10.11924/j.issn.1000-6850.2013-2549

大麻ISSR反应体系的优化与引物的初步筛选

张利国张效霏宋宪友房郁妍郑楠

收稿日期:2013-09-27 修回日期:2013-11-04 出版日期:2014-04-25 发布日期:2014-04-25
基金资助:
黑龙江省自然科学基金项目“大麻种质资源的细胞遗传学研究与ISSR分析”(QC2012C115)；哈尔滨市青年科技创新人才项目“大麻种质资源的遗传多样性研究”(2013RFQYJ014)。

Optimization of ISSR Reaction System of Hemp and Elementary Screening of Primers

Received:2013-09-27 Revised:2013-11-04 Online:2014-04-25 Published:2014-04-25

摘要/Abstract

摘要： 为探寻出更适宜大麻的ISSR反应体系，用以研究大麻的遗传多样性，利用梯度试验对dNTP、Ta qDNA聚合酶和引物的浓度，预变性时间和退火温度5个因素进行优化，从而建立了适合大麻的ISSR-PCR反应体系。结果表明，在25μL体系中，dNTP 200μmol/L，模板1 ng/μL，引物200 nmol/L，聚合酶1 U (2.5 pmol/μL)；预变性3 min (94℃)，退火温度48℃，为大麻最佳反应体系；研究利用优化后的ISSR反应体系，在30条引物中，筛选出8条适合大麻的ISSR引物，体系具有较好的稳定性和重复性。

关键词: 候选基因, 候选基因

Abstract: The five factors affected ISSR reaction, including dNTP, TaqDNA polymerse, concentrations of primer, predegeneration time, annealing temperature etc., were tested to optimize ISSR amplification system by single factor experiment method for finding genetic diversity of hemp. The results showed that: the optimum reaction system in 25μL volume was dNTP 200μmol/L, template 1 ng/μL, primer 200 nmol/L, Taq DNA polymerase 1 U (2.5 pmol/μL), predegeneration time 3 min (94℃), annealing temperature 48℃. Moreover, 8 primers in 30 primers were screened out through the above stable reaction system. The repeatable and stable system was completely suitable to analyze the hemp genetic information.

张利国张效霏宋宪友房郁妍郑楠. 大麻ISSR反应体系的优化与引物的初步筛选[J]. 中国农学通报, 2014, 30(12): 105-109.

参考文献

[1] 关凤芝.大麻遗传育种与栽培技术[M].哈尔滨:黑龙江人民出版社,2010.
[2] 张利国,关凤芝,吴广文,等. 基于核型与 RAPD 标记的大麻地方品种遗传多样性分析[J].中国麻业科学,2009,31(3):169-172.
[3] Hong S, Clarke R C. Taxonomic studies of Cannabis in China[J]. Journal of the International Hemp Association, 1996,3(2):55-60.
[4] Wibowo A C, Mohanty A K, Misra M, et al. Chopped industrial hemp fiber reinforced cellulosic plastic biocomposites: thermomechanical and morphological properties[J]. Industrial & engineering chemistry research,2004,43(16):4883-4888.
[5] Faeti V, Mandolino G, Ranalli P. Genetic diversity of Cannabis sativa germplasm based on RAPD markers[J]. Plant Breeding, 1996, 115(5):367-370.
[6] Nagaoka T, Ogihara Y. Applicability of inter- simple sequence repeat polymorphisms in wheat for use as DNA markers in comparison to RFLP and RAPD markers[J]. Theoretical and applied genetics,1997,94(5):597-602.
[7] Qian W, Ge S, Hong D Y. Genetic variation within and among populations of a wild rice Oryza granulata from China detected by RAPD and ISSR markers[J]. Theoretical and Applied Genetics, 2001,102(2-3):440-449.
[8] Wu C J, Cheng Z Q, Huang X Q, et al. Genetic diversity among and within populations of Oryza granulata from Yunnan of China revealed by RAPD and ISSR markers: implications for conservation of the endangered species[J]. Plant Science,2004,167 (1):35-42.
[9] Zietkiewicz E, Rafalski A, Labuda D. Genome fingerprinting by simple sequence repeat (SSR)-anchored polymerase chain reaction amplification[J]. Genomics,1994,20(2):176-183.
[10] Bornet B, Branchard M. Nonanchored inter simple sequence repeat (ISSR) markers: reproducible and specific tools for genome fingerprinting[J]. Plant molecular biology reporter,2001,19(3):209- 215.
[11] Shang Y, Zhang W H, Pei K Q, et al. Population genetic structure of a dominant desert tree, Haloxylonammodendron (Chenopodiaceae) in the Southeast Gurbantunggut desert detected by RAPD and ISSR markers[J].Acta Botanica Sinica,2004,46(6):675-681.
[12] 张卿哲,孟金贵,张应华,等.用正交设计法优化辣椒 ISSR-PCR 反应体系[J].西南农业学报,2011,24(3):1034-1038.
[13] 邱天.小麦异源多倍体中亲本基因组相互作用产生快速基因组变异的 ISSR 标记分析[J].东北师大学报:自然科学版,2011,2(27.): 124-128.
[14] 郭计华.基于 ISSR 分析 28 份香蕉种质的基因组 DNA 多样性[J].基因组学与应用生物学,2012,31(5):492-497.
[15] 陈名红,李玉,刘多,等.百合属 ISSR 反应条件优化的研究[J].中国农学通报,2013,29(04):118-122.
[16] 卢江.随机放大多态性DNA.[J].植物学报,1993,35 （增刊） :119-127.

大麻ISSR反应体系的优化与引物的初步筛选

Optimization of ISSR Reaction System of Hemp and Elementary Screening of Primers

PDF (PC)

可视化

摘要/Abstract

引用本文

使用本文

参考文献

相关文章 4

编辑推荐

Metrics

本文评价

关于本刊

作者中心

审者中心

在线期刊

联系我们

[1]	刘丽元,周靖航,张梦华,魏趁,王丹,方季青,李金芝,王爱芳,黄锡霞. 新疆褐牛产奶性状候选基因SNPs检测及其遗传效应分析[J]. 中国农学通报, 2017, 33(20): 113-123.
[2]	桑雷孙世坤陈冬金陈岩锋谢喜平. 生殖激素及其受体作为家畜产仔数候选基因的研究进展[J]. 中国农学通报, 2012, 28(26): 1-6.
[3]	张冬杰,刘娣,杨国伟. 野家杂交猪F1代群体8个繁殖功能基因的多态性分析[J]. 中国农学通报, 2009, 25(4): 10-13.
[4]	杨泽峰,张峰,顾世梁,李韬,徐辰武. 利用比较分子遗传学鉴定小麦穗发芽基因的研究进展[J]. 中国农学通报, 2004, 20(5): 50-50.