甜菜TRAP-PCR反应体系的建立及引物开发策略

doi:10.11924/j.issn.1000-6850.casb15080024

中国农学通报 ›› 2016, Vol. 32 ›› Issue (6): 96-101.doi: 10.11924/j.issn.1000-6850.casb15080024

所属专题：园艺

甜菜TRAP-PCR反应体系的建立及引物开发策略

吴则东^1,2,3,刘乃新^1,2,3,秦浩东¹,倪洪涛¹,马龙彪^1,2

(¹黑龙江省普通高校甜菜遗传育种重点实验室/黑龙江大学,哈尔滨 150080；²黑龙江大学农作物研究院,哈尔滨 150080；³农业部糖料产品质量安全风险评估实验室/中国农业科学院甜菜研究所,哈尔滨 150080）

收稿日期:2015-08-05 修回日期:2015-12-24 接受日期:2015-09-18 出版日期:2016-03-07 发布日期:2016-03-07
通讯作者: 马龙彪
基金资助:
甜菜现代产业技术体系建设项目“甜菜丰产抗病种质创新及新品种选育”(CARS-210104-01)。

Establishment of TRAP-PCR Reaction System and Development Strategy of Primers for Beet

Wu Zedong^1,2,3, Liu Naixin^1,2,3, Qin Haodong¹, Ni Hongtao¹, Ma Longbiao^1,2

(¹Key Laboratory of Sugar Beet Genetic Breeding/ Heilongjiang University, Harbin 150080; ²Crop Academy of Heilongjiang University, Harbin 150080;³Laboratory of Quality & Safety Risk Assessment for Sugar Crops Products, Ministry of Agriculture, P.R.China/Sugar Beet Research Institute of Chinese Academy of Agricultural Sciences, Harbin 150080)

Received:2015-08-05 Revised:2015-12-24 Accepted:2015-09-18 Online:2016-03-07 Published:2016-03-07

摘要/Abstract

摘要： 以糖用甜菜为材料，利用单因素实验探讨甜菜TRAP-PCR反应体系中dNTPs、引物、模板以及DNA聚合酶等4个因素的浓度变化对扩增结果的影响，最终建立糖用甜菜的最佳TRAP-PCR反应体系。结果表明，在20 μL反应体系中，甜菜TRAP-PCR最优反应体系包含2.0 μL的10×PCR buffer（含Mg²⁺）、10 ng的模板DNA、0.75 U的Taqase、10 μmol/L固定引物及随机引物各0.8 μL以及0.5 μL的dNTPs (2.5 mmol/L each)。并建立了一种快速开发甜菜TRAP-PCR引物的方法，利用优化后的程序对15份糖甜菜品种进行扩增。结果表明，该体系扩增结果稳定、条带清晰，部分引物多态性丰富，可用于糖用甜菜品种指纹图谱的构建以及其他分子生物学领域的研究。

关键词: 光波, 光波, 光强, 昆虫趋光性, 农业害虫, 趋光节律

Abstract: With sugar beet as the material, the influence of concentration changes of the four factors of dNTPs, primer, template and DNA polymerase in the beet TRAP-PCR reaction system on the amplification results was discussed with single-factor experiment, and finally the optimal TRAP-PCR reaction system of sugar beet was established. The results showed that in the 20 μL reaction system, the beet TRAP-PCR optimal reaction system contained 2.0 μL of 10×PCR buffer (including Mg²), 10 ng of template DNA, 0.75 U of Taqase, respectively 0.8 μL of 10 μmol/L fixed primer and random primer, and 0.5 μL of dNTPs (2.5 mmol/L each). In addition, a method for rapid development of beet TRAP-PCR primer was proposed, 15 varieties of sugar beer were amplified with the optimized program. The results show that the system has stable amplification results, clear bans and abundant polymorphism of partial primer, and can be used for the construction of sugar beet variety fingerprint and research in other fields of molecular biology.

中图分类号:

S566.3

吴则东,刘乃新,秦浩东,倪洪涛,马龙彪. 甜菜TRAP-PCR反应体系的建立及引物开发策略[J]. 中国农学通报, 2016, 32(6): 96-101.

Wu Zedong,Liu Naixin,Qin Haodong,Ni Hongtao and Ma Longbiao. Establishment of TRAP-PCR Reaction System and Development Strategy of Primers for Beet[J]. Chinese Agricultural Science Bulletin, 2016, 32(6): 96-101.

参考文献

[1] Dohm J C, Minoche A E, Holtgr?we D, et al. The genome of the recently domesticated crop plant sugar beet (Beta vulgaris)[J]. Nature,2014,505:546-549.
[2] Halldén C, Hjerdin A, Rading I, et al. A high density RFLP linkage map of sugar beet[J]. Genome.1996,39(4):634-645.
[3] Schondelmaier J, Steinrücken G, Jung C. Integration of AFLP markers into a linkage map of sugar beet (Beta vulgaris L.)[J].Plant breeding,1996,115(4):231-237.
[4] Grimmer M, Trybush S, Hanley S, et al. An anchored linkage map for sugar beet based on AFLP, SNP and RAPD markers and QTL mapping of a new source of resistance to Beet necrotic yellow vein virus[J].Theoretical and Applied Genetics,2007,114(7):1151-1160.
[5] Barzen E, Mechelke W, Ritter E, et al. An extended map of the sugar beet genome containing RFLP and RAPD loci[J]. TAG Theoretical and Applied Genetics,1995,90(2):189-193.
[6] Cureton A, Burns M, Ford‐Lloyd B, et al. Development of simple sequence repeat (SSR) markers for the assessment of gene flow between sea beet (Beta vulgaris ssp. maritima) populations[J]. Molecular Ecology Notes,2002,2(4):402-403.
[7] Barzen E, Mechelke W, Ritter E, et al. RFLP markers for sugar beet breeding: chromosomal linkage maps and location of major genes for rhizomania resistance, monogermy and hypocotyl colour[J]. The Plant Journal,2005,2(4):601-611.
[8] Gidner S, Lennefors B L, Nilsson N O, et al. QTL mapping of BNYVV resistance from the WB41 source in sugar beet[J].Genome,2005,48(2):279-285.
[9] De Riek J, Calsyn E, Everaert I, et al. AFLP based alternatives for the assessment of distinctness, uniformity and stability of sugar beet varieties[J]. Theoretical and Applied Genetics,2001,103(8):1254-1265.
[10] 邓俭英,刘忠,康德贤,等.RFLP分子标记及其在蔬菜研究中的应用[J].分子植物育种,2005,3(2):245-248.
[11] 李珊,赵桂仿.AFLP分子标记及其应用[J].西北植物学报,2003,23 (5):830-836.
[12] 安贤惠,李联泰.RAPD标记在植物种质资源研究中的应用[J].西北农业学报,2000,9(3):126-130.
[13] Hu J, Vick B A. Target region amplification polymorphism: a novel marker technique for plant genotyping[J]. Plant Molecular Biology Reporter,2003,21(3):289-294.
[14] Liu Z, Anderson J, Hu J, et al. A wheat intervarietal genetic linkage map based on microsatellite and target region amplified polymorphism markers and its utility for detecting quantitative trait loci[J]. Theoretical and Applied Genetics,2005,111(4):782-794.
[15] 金梦阳,刘列钊,付福友,等.甘蓝型油菜SRAP、SSR、AFLP和TRAP标记遗传图谱构建[J].分子植物育种,2006,4(4):520-526.
[16] Alwala S, Suman A, Arro J A, et al. Target region amplification polymorphism (TRAP) for assessing genetic diversity in sugarcane germplasm collections[J]. Crop Science,2006,46(1):448-455.
[17] 郭春芳,唐玉海,孙云,等.11个茶树品种遗传多样性的ISSR和TRAP比较分析[J].中国农学通报,2008,24(1):340-346.
[18] Chen J, Hu J, Vick B A, et al. Molecular mapping of a nuclear male-sterility gene in sunflower (Helianthus annuus L.) using TRAP and SSR markers[J]. Theoretical and Applied Genetics,2006,113(1):122-127.
[19] 吴则东,王茂芊,胡珅,等.一种适用于PCR扩增的甜菜干种子DNA快速提取方法的研究[J].中国农学通报,2013,29(30):69-72.
[20] 王凤格,赵久然,郭景伦,等.一种改进的玉米SSR标记的PAGE/快速银染检测新方法[J].农业生物技术学报,2004,12(5):606-607.
[21] 张丽群,成浩,王丽鸳,等.茶树TRAP反应体系的建立及正交设计优化[J].中国农学通报,2012,28(28):220-226.
[22] 王亚楠,韩莹琰,范双喜.叶用莴苣TRAP反应体系的建立[J].中国农学通报,2014,30(28):99-104.
[23] 任民,王志德,贾兴华,等.开发TRAP标记的新策略[J].中国烟草科学,2009,30(S1):25-31.
[24] Sen A, Alikamanoglu S. Analysis of drought-tolerant sugar beet (Beta vulgaris L.) mutants induced with gamma radiation using SDS-PAGE and ISSR markers[J].Mutation Research/Fundamental and Molecular Mechanisms of Mutagenesis,2012,738:38-44.
[25] Izzatullayeva V, Akparov Z, Babayeva S, et al. Efficiency of using RAPD and ISSR markers in evaluation of genetic diversity in sugar beet[J].Turkish Journal of Biology,2014,38(4):429-438.
[26] Grimmer M, Trybush S, Hanley S, et al. An anchored linkage map for sugar beet based on AFLP, SNP and RAPD markers and QTL mapping of a new source of resistance to Beet necrotic yellow vein virus[J]. TAG Theoretical and Applied Genetics,2007,114(7):1151-1160.
[27] Li J, Schulz B, Stich B. Population structure and genetic diversity in elite sugar beet germplasm investigated with SSR markers[J]. Euphytica,2010,175(1):35-42.

甜菜TRAP-PCR反应体系的建立及引物开发策略

Establishment of TRAP-PCR Reaction System and Development Strategy of Primers for Beet

PDF (PC)

可视化

摘要/Abstract

引用本文

使用本文

参考文献

相关文章 12

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]	吕冬煦, 倪海瑞, 曾彦达. 光照强度对水稻HS叶片失绿的影响[J]. 中国农学通报, 2020, 36(20): 23-27.
[2]	徐练,文礼章,易倩,胡亮,彭云鹏. 不同光波、光源距离及环境温度对异色瓢虫成虫趋光行为的影响[J]. 中国农学通报, 2016, 32(10): 106-113.
[3]	蒋菊芳,赵福年,张宇林,任丽雯,杨永龙. 干旱胁迫下春玉米叶片对光强和CO₂浓度的反应[J]. 中国农学通报, 2015, 31(36): 43-48.
[4]	张磊. 3种专用型甘薯光合光响应曲线及其模型拟合[J]. 中国农学通报, 2015, 31(15): 71-77.
[5]	杨彩宏冯莉田兴山张泰劼高家东岳茂峰崔烨. 不同秸杆水提液在不同光照强度下对马唐萌发与生长的影响[J]. 中国农学通报, 2014, 30(4): 270-274.
[6]	杨洪璋文礼章易倩许浩. 光波和光强对几种重要农业害虫趋光性的影响[J]. 中国农学通报, 2014, 30(25): 279-285.
[7]	陈坤文礼章龚碧涯杨洪璋李小一许浩. 重要气象因子对农田灯光诱虫效率的影响[J]. 中国农学通报, 2013, 29(4): 44-50.
[8]	李季春. 内蒙古羊草草原主要建群草种的光合参数[J]. 中国农学通报, 2010, 26(1): 227-230.
[9]	闫新房,丁林波,丁义,何松林. LED在植物组织培养中的应用[J]. 中国农学通报, 2009, 25(12): 42-45.
[10]	李佩华,,彭徐. 马铃薯遮光处理的效应研究[J]. 中国农学通报, 2007, 23(4): 220-220.
[11]	卜登攀,李树聪,张瑞超,周凌云,于建国,魏宏阳,王加启. FAST指数在复原乳区分中的应用初探[J]. 中国农学通报, 2006, 22(11): 1-1.
[12]	吴瑛,袁守亮. 转基因生物农药的安全性评价[J]. 中国农学通报, 2005, 21(8): 370-370.