摘要
硼是植物生长发育的必需营养元素,在植物体内具有重要的生理功能。然而,土壤缺硼是世界范围内重要的农业生产问题,成为限制多种作物产量的主要因子。甘蓝型油菜是世界上主要的油料作物之一,是需硼较高的物种,且对缺硼敏感。但是,不同甘蓝型油菜品种对低硼胁迫存在极显著的基因型差异。本研究利用筛选出的硼高效(抗低硼胁迫)品种青油10号和硼低效(对低硼胁迫敏感)品种Westar10构建的QW DH群体,对甘蓝型油菜硼高效相关性状进行了QTL分析、同时构建了硼高效主效QTL的近等基因系,获得的主要结果如下：
1.甘蓝型油菜硼高效相关性状QTL的检测
利用420个SSR标记和153个AFLP标记构建了QW DH遗传连锁图谱,该图谱覆盖了1923.15cM的遗传距离,平均标记密度为0.30marker/cM,相邻分子间的平均距离为3.35cM。通过三次独立重复营养液培养试验调查QW DH群体及亲本材料在正常硼(25μM B)和低硼(0.25μM B)水平下的表型,选取根长增加量、地上部干重、根干重、地上部硼含量、根硼含量、地上部硼累积量、根硼累积量和硼效率系数共8个性状为硼高效相关性状。利用QW DH图谱对这8个硼高效相关性状进行QTL扫描,共检测到71个QTLs,其中,包括正常硼水平下30个QTLs,低硼水平下35个QTLs和硼效率系数6个QTLs,单个QTL的表型贡献率范围为4.58%-48.34%。所检测到的QTLs分布在QW DH遗传连锁图谱的18条连锁群上,其中在A3,A4,A5和C4连锁群上存在着QTL簇。位于A3连锁群58.8-82.5cM区间的QTL簇,QTL-cluster-A3a,包含低硼水平下调控根长增加量、地上部干重、根干重、地上部硼累积量、根中硼累积量和硼效率系数的QTLs,贡献率为14.80%-48.34%,被认为是控制甘蓝型油菜硼高效的主效QTL位点。
2.甘蓝型油菜高密度遗传连锁图谱的构建及硼高效相关性状QTL的检测
基于甘蓝型油菜60K SNP:芯片技术,在上述QW DH群体中获得11,754个SNP标记,分为1,346个SNP-bin。与上述QW DH遗传连锁图谱整合,获得一张包含1,398个标记的高密度遗传连锁图谱,覆盖遗传距离为2139.34cM,平均分子标记密度为0.65marker/cM,相邻分子间的平均距离为1.53cM。根据两图谱上共有的标记,高密度图谱与原图谱间具有较好的线性关系。利用高密度遗传连锁图谱重新检查硼高效相关性状的QTL,共获得75个QTLs,正常硼和低硼水平下分别为30和41个QTLs,硼效率系数4个QTLs。单个QTL的表型贡献率范围为7.26%-31.56%。其中,有34个QTLs可在两个图谱中重复检测到。另外,在A3,A4和C4连锁群上同样存在QTL簇的分布,这些QTL簇与先前QW DH图谱检测的QTL簇位于染色体的相同位置。
3.甘蓝型油菜三个硼高效QTL簇近等基因系的构建
根据QW DH群体的硼高效QTL检查结果,将QTL-cluster-A3a, QTL-cluster-A4和QTL-cluster-A5作为目标QTLs。以硼低效亲本Westar10为轮回亲本,通过连续不断回交,构建其近等基因系。通过每一回交世代单株抗低硼胁迫表型鉴定、高世代回交单株目标QTLs簇的前背景和后背景基因型分析,最终获得6个含有不同硼高效目标QTL簇的BC3F1单株自交发展的BC4F1株系。对这6个株系进行抗低硼表型鉴定,均表现出明显的抗缺硼表型分离。其中,以同时含有3个目标QTLs簇的BC4F1-3株系的表型分离最为明显。在含有硼高效主效QTL-cluster-A3a的BC4F1-2株系中,筛选到一个在QTL区间基因型为杂合、其它区间为Westar10纯和的单株,进一步通过自交获得该单株的BC4F2群体。在该群体中,筛选到含有不同目标QTL片段的染色体代换系,根据后代表型测验,将QTL-cluster-A3a锁定在A3连锁群的CNU384-BnGMS436区间。
4.甘蓝型油菜地上部矿质元素QTL的分析
调查QW DH群体在正常硼和低硼水平下的地上部矿质元素的含量。与正常硼相比,低硼降低了地上部K和Mn的含量,增加了P,Fe和Zn的含量,对于Ca,Mg和Cu的含量影响不大。矿质元素在不同的硼水平下存在着显著的正相关或负相关。利用QW DH遗传连锁图,对地上部矿质元素含量进行QTL分析。分别检测到控制甘蓝型油菜地上部大中量元素和微量元素的QTLs为28和27个,表型贡献率为10.44%-30.97%。控制不同矿质元素含量的QTLs在QW DH遗传连锁图谱上存在共定位的现象。
Boron (B) is an essentical element for the growth and development of plants, which has the important physiological functions in plants. However, B deficiency in soils is a major agricultural problem worldwide, becoming a main limiting factor for the production of many crops. Oilseed rape (Brassica napus L.) is one main oil crop in the world, which is a high B demand species and is extremely sensitive to B deficiency. Significant genotypic vatiation was existed among the different B. napus cultivars in response to B deficiency. In this study, quantitative trait locus (QTL) for B-efficiency traits in B. napus was analyzed using the QW DH population derived from a cross between B-efficienct parent Qingyou10and B-inefficient parent Westar10, and near-isgenic lines (NILs) for major effect QTLs for B efficiency were also developped. The main results are as follows:
1. QTL analysis for B-efficiency traits in B. napus
The QW DH genetic map was constructed using420simple sequence repeat (SSR) and153amplified fragment length polymorphism (AFLP) markers. The total length of this map was1923.15cM with an average marker density of0.30marker/cM and an average interval of3.35cM between adjacent markers. Phenotypes of QW DH population and their partental lines were investigated under normal B (25μM B) and low B (0.25μM B) levels by three independent solution culture trials. A total of eight traits were selected as the B-efficiency traits, including increment of primary root length (IPRL), shoot dry weight (SDW), root dry weight (RDW), shoot B concentration (SBC), root B concentration (RBC), shoot B accumulation (SBA), root B accumulation (RBA) and B efficiency coefficient (BEC). The QTLs for these eight traits were analyzed using the QW DH genetic map. A total of71QTLs were obstained, including30under normal B level,35under low B level and6for BEC, which explained single the phenotypic variation ranged from4.58%to48.34%. The QTLs distributed on eighteen linkage groups in QW DH genetic map. Moreover, the QTL clusters were observed on A3, A4, A5and C4linkage groups. QTL-cluster-A3a located in the interval of58.8-82.5cM on A3, including the QTLs for IPRL, SDW, RDW, SBA, RBA at low B level and BEC whose phenotypic varation ranged from14.80%-48.34%, was deemed as the major QTL for B efficiency of B. napus.
2. Construction of high density genetic map and QTL mapping for B-efficiency traits
Based on the Brassica60K SNP BeadChip Array, a total of11,754single nucleotide polymorphism (SNP) markers were obstained in the QW DH population, classified into the1,346bins. The high density genetic map was constructed using these SNP-bins and the primary markers in the QW DH genetic map. The final high density genetic map contained1,398markers, covered2139.34cM, with an average marker density of0.65marker/cM and an average distance between adjacent markers of1.53cM. Linear relationship was observed between two genetic maps based on the commom marker. The QTL for B-efficiency traits was reanalyzed using the high density genetic map. In total,75QTLs were detected for associated with B efficiency. Of them,30QTLs were detected under normal B level,41QTLs were detected under low B level and4QTLs for BEC, respectively. Each QTL accounted for7.26%-31.56%of phenotypic variation. Among these QTLs,34were common detected across QW DH and high density genetic maps. Moreover, the QTL clusters on A3, A4and C4were also observed in high density genetic map, which located on the same chromosome regions with those observed in QW DH genetic map.
3. Construction near isogenic lines of three QTL clusters for B efficiency in B. napus
Based on the QTL results in QW DH population, QTL-cluster-A3a, QTL-cluster-A4and QTL-cluster-A5were selected as the target QTLs for construction the NIL. Backcross populations were constructed using B-inefficienct parent Westar10as recurrent parent. Every progenies were screened by the phenotypc investigation response to B deficiency and foreground and bachgroud analysis for high progenies, finlly, six BC4F1lines derived from a cross between BC3F1contained the different target QTLs and Westar10were used for investigation the phenotypic varation response to B deficiency. BC4F1.3, including three target QTLs simultaneously, showed the most widely variations among six lines. Substitution mapping revealed that the major QTL, QTL-cluster-A3a, was mapped in the interval between CNU384-BnGMS436.
4. QTL analysis for mineral concentraions in shoot of B. napus
The mineral concentrations in shoot of QW DH population were investigated under normal B and low B levels. Compared with normal B, the concentrations of K and Mn were increased by B deficiency, however, the concentrations of P, Fe and Zn were decreased. The concentrations of Ca, Mg and Cu were not affected by B deficiency. Significant positive or negative relationships were observed among most of mineral concentrations. QTL for mineral concentrations was analyzed using QW DH genetic map. A total of28and27QTLs for macro-element concentrations and micro-element concentrations were detected, resceptively, explaining10.44%-30.97%of phenotypic variation. The co-locations of QTLs for different mineral concentrations were observed in QW DH genetic map.
引文
1.丁广大,刘佳,石磊,徐芳森.植物离子组学：植物营养研究的新方向.植物营养与肥料学报,2010,16(2)：479-484.
2.杜昌文,王运华,徐芳森,王火焰.不同硼效率甘蓝型油菜品种中硼的形态及其相互关系.植物营养与肥料学报,2002,8(1)：105-109.
3.方宣钧,吴为人,唐纪良.作物DNA标记辅助育种.北京：科学出版社,2000.
4.何建新.植物对硼吸收转运机理的研究进展.中国沙漠,2008,28(2)：266-273.
5.李赢赢.甘蓝型油菜硼高效遗传连锁图的构建及相关QTL定位.[硕士学位论文].武汉：华中农大学图书馆,2011.
6.刘佳.甘蓝型油菜硼营养及离子组遗传学研究.[博士学位论文].武汉：华中农大学图书馆,2009.
7.刘武定,皮美美,吴礼树.棉花硼、钾营养相互关系的研究.土壤学报,1987,24(1)：43-50.
8.刘铮,朱其清,唐丽华.土壤中硼的含量和分布的规律性.土壤学报,1989,26(4)：353-361.
9.陆景陵.植物营养学(上册).北京：中国农业大学出版社,2003.
10.马智宏,关军锋.钙,硼水平对苹果花粉萌发和花粉管生长的影响.河北农业科学,1999,3(2)：1-4.
11.年夫照,彭慧元,徐芳森,石磊,王运华.不同硼效率甘蓝型油菜苗期对硼镁营养的反应.植物营养与肥料学报,2004,10(5)：511-515.
12.潘存红,王子斌,马玉银,殷跃军,张亚芳,左示敏,陈宗祥,潘学彪.InDel和SNP标记在水稻图位克隆中的应用.中国水稻科学,2007,21(5)：447-453.
13.潘媛.不同油菜品种硼吸收利用差异与细胞壁组分关系的研究.[硕士学位论文].武汉：华中农大学图书馆,2010.
14.沈振国,沈康,张秀省.油菜幼苗对硼的吸收与运转及钙的影响.植物学通报,1992,9(4)：33-37.
15.石磊.甘蓝型油菜硼高效的生理基础及硼高效基因的定位.[博士学位论文].武汉：华中农业大学图书馆,2005.
16.石磊,杨玉华,徐芳森,王运华.硼对作物细胞膜功能影响的研究进展.华中农业大学学报,2002,21：395-400.
17.石磊,年夫照,赵华,徐芳森,孟金陵,王运华.7个甘蓝型油菜品种对硼胁迫反应的差异.中国油料作物学报,2004,26(1)：48-51.
18.孙进华.甘蓝型油菜硼转运子基因的克隆、表达和分子进化.[博士学位论文].武汉：华中农业大学图书馆,2011.
19.王火焰,王运华.不同硼效率甘蓝型油菜品种悬浮细胞的硼钙营养效应.植物营养与肥料学报,2002,100-104.
20.王运华,兰莲芳.甘蓝型油菜品种对缺硼敏感性差异的研究(Ⅰ,Ⅱ,Ⅲ).华中农业大学学报,1995,21：71-84
21.熊汉峰,刘武定,皮美美.硼氮及其配合对油菜苗期养分吸收的影响.土壤通报,1995,26：84-86.
22.徐芳森,王运华,李建春.甘蓝型油菜硼营养高效在F_1代的遗传研究.植物营养与肥料学报,1998a,4(3),305-310.
23.徐芳森,王运华,李建春.甘蓝型油菜不同硼效率品种对缺硼反应的研究.华中农业大学学报,1998b,17(1),59-64.
24.薛建明,杨玉爱.不同油菜基因型对缺硼反应的差异.浙江农业大学学报,1994,20,422-426.
25.杨锦鹏.利用TN群体定位甘蓝型油菜硼高效QTLs的研究.[硕士学位论文].武汉：华中农大学图书馆,2007.
26.杨露.甘蓝型油菜不同硼效率品种硼吸收转运差异的机理研究.[硕士学位论文].武汉：华中农大学图书馆,2012.
27.喻敏,褚海燕,吴礼树,王运华.甘蓝型油菜不同硼效率基因型对硼的吸收、分配的影响.中国油料作物学报,1999,21(1),49-53.
28.曾丽霞,艾应伟.硼对花粉生物化学功能影响的研究进展.中国土壤与肥料,2008,(3),9-11.
29.张秀省,沈振国.硼对油菜花器官发育和结实性的影响.土壤学报,1994,31(2)：146-162.
30.赵华.甘蓝型油菜硼高效近等基因系的构建及硼高效基因BnBE2的定位.[博士学位论文].武汉：华中农业大学图书馆,2008.
31.赵竹青,王运华,吴礼树.缺硼对黄瓜生长素代谢的影响.华中农业大学学报,1998,17(3),32-36.
32.朱军.运用混合线性模型定位复杂数量性状基因的方法.浙江大学学报(自然科学版),1999,33(3),111-119.
33. Alpert KB, Tanksley SD. High-resolution mapping and isolation of a yeast artificial chromosome contig containing fw2.2:a major fruit weight quantitative trait locus in tomato. Proc Natl Acad Sci,1996,93:15503-15507.
34. Arabidopsis Genome Initiative. Analysis of the genome sequence of the flowering plant Arabidopsis thaliana. Nature,2000,408:796-815.
35. Bancroft I, Morgan C, Fraser F, Higgins J, Wells R, Clissold L, Baker D, Long Y, Meng J, Wang X, Liu S, Trick M. Dissecting the genome of the polyploid crop oilseed rape by transcriptome sequencing. Nat Biotechnol,2011,29:762-766.
36. Brown PH, Bellaloui N, Hu H, Dandekar A. Transgenically enhanced sorbitol synthesis facilitates phloem boron transport and increases tolerance of tobacco to boron deficiency. Plant Physiol,1999,119:17-20.
37. Brown PH, Bellaloui N, Wimmer MA, Bassil ES, Ruiz J, Hu H, Pfeffer H, Dannel F, Romheld V. Boron in plant biology. Plant Biol,2002,4:205-223.
38. Brown PH, Shelp BJ. Boron mobility in plants. Plant Soil,1997,193:85-101.
39. Cakmak I, Kurz H, Marschner H. Short-term effects of boron, germanium and high light intensity on membrane permeability in boron deficient leaves of sunflower. Physiol Plant,1995,95:11-18.
40. Canon P, Aquea F, Rodr i guez-Hoces GA, Arce-Johnson P. Functional characterization of Citrus macrophylla BOR1 as a boron transporter. Physiol Plant, 2013,149:329-339.
41. Chatterjee C, Sinha P, Agarwala SC. Iteractive effect of boron and phosphorus on growth and metabolism of maize grown in refined sand. Can J Plant Sci, 1990, 70: 455-460.
42. Cheung F, Trick M, Drou N, Lim YP, Park J, Kwon S, Kim J, Scott R, Pires JC, Paterson A H, Town C, Bancroft I. Comparative analysis between homoeologous genome segments of Brassica napus and its progenitor species reveals extensive sequence-level divergence. Plant Cell, 2009, 21:1912-1928.
43. Chormova D, Messenger DJ, Fry SC. Boron bridging of rhamnogalacturonan-II, monitored by gel electrophoresis, occurs during polysaccharide synthesis and secretion but not post-secretion. Plant J, 2014, 77:534-546.
44. Clemens S. Molecular mechanisms of plant metal tolerance and homeostasis. Planta, 2001,212:475-486.
45. Dannel F, Pfeffer H, Romheld V. Characterization of root boron pools, boron uptake and boron translocation in sunflower using the stable isotopes 10B and 11B. Funct Plant Biol,2000,27:397-405.
46. Dell B, Huang L. Physiological response of plants to low boron. Plant Soill,1997, 193:103-120.
47. Dordas C, Brown PH. Permeability of boric acid across lipid bilayers and factors affecting It. JMembr Biol, 2000, 175:95-105.
48. Dordas C, Chrispeels MJ, Brown PH. Permeability and channel-mediated transport of boric acid across membrane vesicles isolated from squash roots. Plant Physiol, 2000, 124:1349-1362.
49. Eulgem T, Rushton PJ, Robatzek S, Somssich IE. The WRKY superfamily of plant transcription factors. Trends Plant Sci, 2000, 5:199-206.
50. Eulgem T, Somssich IE. Networks of WRKY transcription factors in defense signaling. Curr Opin Plant Biol,2007,10:366-371.
51. Ferreira ME, Satagopan J, Yandell BS, Williams PH, Osborn TC. Mapping loci controlling vernalization requirement and flowering time in Brassica napus. Theor Appl Genet,1995,90:727-732.
52. Frary A, Nesbitt TC, Frary A, Grandillo S, Knaap E, Cong B, Liu J, Meller J, Elber R, Alpert KB, Tanksley SD. fw2.2:a quantitative trait locus key to the evolution of tomato fruit size. Science,2000,289:85-88.
53. Gunes A, Alpaslan M. Boron uptake and toxicity in maize genotypes in relation to boron and phosphorus supply. J Plant Nutr,2000,23:541-550.
54. Hasan M, Seyis F, Badani AG, Pons-Kuhnemann J, Friedt W, Luhs W, Snowdon RJ. Analysis of genetic diversity in the Brassica napus L. gene pool using SSR markers. Genet Resour Crop Evol, 2006, 53:793-802.
55. Hoagland DR, Arnon DI. The water-culture method for growing plants without soil. Circ Calif Agric Exp Stn,1950,347:32
56. Huang L, Bell RW, Dell B. Boron supply into wheat Triticum aestivum L. cv. Wilgoyne ears whilst still enclosed within leaf sheaths. J Exp Bot,2001,52: 1731-1738.
57. Ishii T, Matsunaga T. Isolation and characterization of a boron-rhamnogalacturonan-II complex from cell walls of sugar beet pulp. Carbohydr Res,1996,284:1-9.
58. Ishii T, Matsunaga T, Hayashi N. Formation of rhamnogalacturonan II-borate dimer in pectin determines cell wall thickness of pumpkin tissue. Plant Physiol, 2001,126: 1698-1705.
59. Iwai H, Hokura A, Oishi M, Chida H, Ishii T, Sakai S, Satoh S. The gene responsible for borate cross-linking of pectin Rhamnogalacturonan-II is required for plant reproductive tissue development and fertilization. Proc Natl Acad Sci, 2006, 103: 16592-16597.
60. Iwai H, Masaoka N, Ishu T, Satoh S. A pectin glucuronyltransferase gene is essential for intercellular attachment in the plant meristem. Proc Natl Acad Sci, 2002, 99: 16319-16324.
61. Kao CH, Zeng ZB, Teasdale RD. Multiple interval mapping for quantitative trait loci. Genetics,1999,152:1203-1216.
62. Kasajima I, Ide Y, Yokota Hirai M, Fujiwara T. WRKY6 is involved in the response to boron deficiency in Arabidopsis thaliana. Physiol Plant, 2010, 139:80-92.
63. Kato Y, Miwa K, Takano J, Wada M, Fujiwara T. Highly boron deficiency-tolerant plants generated by enhanced expression of NIP5;1, a boric acid channel. Plant Cell Physiol, 2009,50:58-66.
64. Kaya C, Tuna AL, Dikilitas M, Ashraf M, Koskeroglu S, Guneri M. Supplementary phosphorus can alleviate boron toxicity in tomato. Sci Hortic, 2009, 121:284-288.
65. Kobayashi M. Matoh,T. Azuma J. Two chains of rhamnogalacturonan II are cross-linked by borate-diol ester bonds in higher plant cell walls. Plant Physiol, 1996, 110:1017-1020.
66. Kosambi DD. The estimation of map distances from recombination values. Ann Eugen,1943,12:172-175.
67. Lahner B, Gong J, Mahmoudian M, Smith EL, Abid KB, Rogers EE, Guerinot ML, Harper JF, Ward JM, Mcintyre L, Schroeder J, Salt DE. Genomic scale profiling of nutrient and trace elements in Arabidopsis thaliana. Nat Biotechnol, 2003,21: 1215-1221.
68. Lander ES, Botstein D. Mapping mendelian factors underlying quantitative traits using RFLP linkage maps. Genetics, 1989, 121:185-199.
69. Leaungthitikanchana S, Fujibe T, Tanaka M, Wang S, Sotta N, Takano J, Fujiwara T. Differential expression of three BOR1 genes corresponding to different genomes in response to boron conditions in hexaploid wheat (Triticum aestivum L.). Plant Cell Physiol,2013,54:1056-1063.
70. Li H, Ye G, Wang J. A modified algorithm for the improvement of composite interval mapping. Genetics, 2007, 175:361-374.
71. Liu L, Qu C, Wittkop B, Yi B, Xiao Y, He Y, Snowdon RJ, Li J. A high-density SNP map for accurate mapping of seed fibre QTL in Brassica napus L. PLoS ONE, 2013, 8:e83052.
72. Marschner P. Marschner's Mineral Nutrition of Higher Plants. Academic Press,2012.
73. Matoh T, Ishigaki K, Mizutani M, Matsunaga W, Takabe K. Boron nutrition of cultured tobacco BY-2 cells I. Requirement for and intracellular localization of boron and selection of cells that tolerate low levels of boron. Plant Cell Physiol, 1992, 33: 1135-1141.
74. Matoh T, Ishigaki K, Ohno K, Azuma J. Isolation and characterization of a boron-polysaccharide complex from radish roots. Plant Cell Physiol, 1993, 34: 639-642.
75. Matoh T, Kawaguchi S, Kobayashi M. Ubiquity of a borate-rhamnogalacturonan II complex in the cell walls of higher plants. Plant Cell Physiol, 1996, 37:636-640.
76. Miwa K, Fujiwara T. Boron transport in plants: co-ordinated regulation of transporters. Ann Bot, 2010,105:1103-1108.
77. Miwa K, Takano J, Fujiwara T. Improvement of seed yields under boron-limiting conditions through overexpression of BOR1, a boron transporter for xylem loading, in Arabidopsis thaliana. Plant J, 2006, 46:1084-1091.
78. Miwa K, Takano J, Omori H, Seki M, Shinozaki K, Fujiwara T. Plants tolerant of high boron levels. Science, 2007, 318:1417-1417.
79. Miwa K, Wakuta S, Takada S, Ide K, Takano J, Naito S, Omori H, Matsunaga T, Fujiwara T.Roles of BOR2, a boron exporter, in cross linking of rhamnogalacturonan II and root elongation under boron limitation in Arabidopsis. Plant Physiol, 2013, 163:1699-1709.
80. Nakagawa Y, Hanaoka H, Kobayashi M, Miyoshi K, Miwa K, Fujiwara T. Cell-type specificity of the expression of OsBOR1, a rice efflux boron transporter gene, is regulated in response to boron availability for efficient boron uptake and xylem loading. Plant Cell, 2007, 19:2624-2635.
81. Noguchi K, Dannel F, Pfeffer H, Romheld V, Hayashi H, Fujiwara T. Defect in root-shoot translocation of boron in Arabidopsis thaliana mutant bor1-1. J Plant Physiol,2000,156:751-755.
82. Noguchi K, Yasumori M, Imai T, Naito S, Matsunaga T, Oda H, Hayashi MC, Fujiwara T. borl-1, an Arabidopsis thaliana mutant that requires a high level of boron. Plant Physiol,1997,115:901-906.
83. O'Neill MA, Eberhard S, Albersheim P, Darvill AG. Requirement of borate cross-linking of cell wall rhamnogalacturonan II for Arabidopsis growth. Science, 2001,294:846-849.
84. O'Neill MA, Ishii T, Albersheim P, Darvill AG. Rhamnogalacturonan II:structure and function of a borate cross-linked cell wall pectic polysaccharide. Annu Rev Plant Biol,2004,55:109-139.
85. O'Neill MA, Warrenfeltz D, Kates K, Pellerin P, Doco T, Darvill AG, Albersheim P. Rhamnogalacturonan-Ⅱ, a pectic polysaccharide in the walls of growing plant cell, forms a dimer that is covalently cross-linked by a borate ester. In vitro conditions for the formation and hydrolysis of the dimer. J Biol Chem,1996,271:22923-22930.
86. Pabst M, Fischl RM, Brecker L, Morelle W, Fauland A, Kofeler H, Altmann F, Leonard R. Rhamnogalacturonan II structure shows variation in the side chains monosaccharide composition and methylation status within and across different plant species. Plant J,2013,76:61-72.
87. Pan Y, Wang Z, Yang L, Wang Z, Shi L, Naran R, Azadi P, Xu F. Differences in cell wall components and allocation of boron to cell walls confer variations in sensitivities of Brassica napus cultivars to boron deficiency. Plant Soil,2012,354: 383-394.
88. Perez-Castro R, Kasai K, Gainza-Cortes F, Ruiz-Lara S, Casaretto.A, Pena-Cort6s H, Tapia J, Fujiwara T, Gonzalez E. VvBORl, the grapevine ortholog of AtBORl, encodes an efflux boron transporter that is differentially expressed throughout reproductive development of Vitis vinifera L. Plant Cell Physiol,2012,53:485-494.
89. Prasad SR, Bagali PG, Shailaja H, Shashidhar HE. Molecular mapping of quantitative trait loci associated with seedling tolerance to salt stress in rice(Oryza sativa L.). Curr Sci,2000,78:162-164.
90. Raven JA. Short-and long-distance transport of boric acid in plants. New Phytol, 1980,84:231-249.
91. Rawson H. The developmental stage during which boron limitation causes sterility in wheat genotypes and the recovery of fertility. Funct Plant Biol,1996,23:709-717.
92. Rodolphe F, Lefort M. A multi-marker model for detecting chromosomal segments displaying QTL activity. Genetics, 1993,134:1277-1288.
93. Salt DE, Baxter I, Lahner B. Ionomics and the study of the plant ionome. Annu Rev Plant Biol,2008,59:709-733.
94. Sax K. The association of size differences with seed-coat pattern and pigmentation in phaseolus vulgaris. Genetics,1923,8:552-560.
95. Schon MK, Novacky A, Blevins DG. Boron induces hyperpolarization of sunflower root cell membranes and increases membrane permeability to K+. Plant Physiol,1990, 93:566-571.
96. Shorrocks VM. The occurrence and correction of boron deficiency. Plant Soil,1997, 193:121-148.
97. Simpson SP. Detection of linkage between quantitative trait loci and restriction fragment length polymorphisms using inbred lines. Theor Appl Genet,1989,77: 815-819.
98. Singh UM, Sareen P, Sengar RS, Kumar A. Plant ionomics:a newer approach to study mineral transport and its regulation. Acta Physiol Plant,2013,35:2641-2653.
99. Stangoulis JCR, Grewal HS, Bell RW, Graham RD. Boron efficiency in oilseed rape: I. Genotypic variation demonstrated in field and pot grown Brassica napus L. and Brassica juncea L. Plant Soil,2000,225:243-251.
100.Stangoulis JCR, Reid RJ, Brown PH, Graham RD. Kinetic analysis of boron transport in Chara. Planta, 2001,213:142-146.
101.Stangoulis JCR, Webb MJ, Graham RD. Boron efficiency in oilseed rape:Ⅱ. Development of a rapid lab-based screening technique. Plant Soil, 2000b, 225: 253-261.
102.Sun J, Shi L, Zhang C, Xu F. Cloning and characterization of boron transporters in Brassica napus. Mol Biol Rep, 2012,39:1963-1973.
103.Sutton T, Baumann U, Hayes J, Collins NC, Shi B, Schnurbusch T, Hay A, Mayo G, Pallotta M,Tester M, Langridge P. Boron-toxicity tolerance in barley arising from efflux transporter amplification. Science,2007,318:1446-1449.
104.Takano J, Miwa K, Fujiwara T. Boron transport mechanisms:collaboration of channels and transporters. Trends Plant Sci,2008,13:451-457.
105.Takano J, Noguchi K, Yasumori M, Kobayashi M, Gajdos Z, Miwa K, Hayashi H, Yoneyama T, Fujiwara T. Arabidopsis boron transporter for xylem loading. Nature, 2002,420:337-340.
106.Takano J, Wada M, Ludewig U, Schaaf G, Wiren N, Fujiwara T. The Arabidopsis major intrinsic protein NIP5;1 is essential for efficient boron uptake and plant development under boron limitation. Plant Cell,2006,18:1498-1509.
107.Tanaka M, Fujiwara T. Physiological roles and transport mechanisms of boron: perspectives from plants. Pflug Arch-Eur J Physiol,2008,456:671-677.
108.Tanaka M, Wallace IS, Takano J, Roberts DM, Fujiwara T. NIP6;1 is a boric acid channel for preferential transport of boron to growing shoot tissues in Arabidopsis. Plant Cell,2008,20:2860-2875.
109.Trick M, Adamski NM, Mugford SG, Jiang C, Febrer M, Uauy C. Combining SNP discovery from next-generation sequencing data with bulked segregant analysis (BSA) to fine-map genes in polyploid wheat. BMC Plant Biol,2012,12:14.
110.Ulloa M, Meredith WR. Genetic linkage map and QTL analysis of agronomic and fiber quality traits in an intraspecific population. J Cotton Sci,2000,4:161-170.
111.Van Ooijen JW. JionMap 4.0:software for the calculation of genetic linkage maps in expermental population. Kyazma BV, Wageningen, Netherlands.
112.Wang G, Romheld V, Li C, Bangerth F. Involvement of auxin and CKs in boron deficiency induced changes in apical dominance of pea plants (Pisum sativum L.). J Plant Physiol,2006,163:591-600.
113.Wang X, Wang H, Wang J, Sun R, Wu J, Liu S, Bai Y, Mun J, Bancroft L, Cheng F, Huang S, Li X, Hua W, Wang J, Wang X, Freeling M, Pires J, Paterson A, Chalhoub B, Wang B, et al. The genome of the mesopolyploid crop species Brassica rapa. Nat Genet,2011,43:1035-1039.
114.Wang Z, Wang Z, Chen S, Shi L, Xu F. Proteomics reveals the adaptability mechanism of Brassica napus to short-term boron deprivation. Plant Soil,2011,347: 195-210.
115. Wang Z, Wang Z, Shi L, Wang L, Xu F. Proteomic alterations of Brassica napus root in response to boron deficiency. Plant Mol Biol,2010,74:265-278.
116.Warington K. The effect of boric acid and borax on the broad bean and certain other plants. Ann Bot,1923,37:629-672.
117.Wimmer MA, Lochnit G, Bassil E, M u hling KH, Goldbach HE. Membrane-associated, boron-interacting proteins isolated by boronate affinity chromatography. Plant Cell Physiol,2009,50:1292-1304.
118.Xu F, Yang Y, Wang Y, Wu L. Boron uptake and retranslocation in cultivars of Brassica napus differing in boron efficiency. In: Goldbach HE, Brown PH, Rerkasem B, Thellier M, Wimmer M., Bell RW eds., Boron in Plant and Animal Nutrition: Springer Press, 2002. 127-135
119.Xu FS, Wang YH, Meng J. Mapping boron efficiency gene(s) in Brassica napus using RFLP and AFLP markers. Plant Breed, 2001,120:319-324.
120.Xue J, Lin M, Bell RW, Graham RD, Yang X, Yang Y. Differential response of oilseed rape (Brassica napus L.) cultivars to low boron supply. Plant Soil,1998,204: 155-163.
121. Yang L, Zhang Q, Dou J, Li L, Guo L, Shi L, Xu F. Characteristics of root boron nutrition confer high boron efficiency in Brassica napus cultivars. Plant Soil,2013, 371:95-104.
122.Yang Y, Lai K, Tai P, Li W. Rates of nucleotide substitution in angiosperm mitochondrial DNA sequences and dates of divergence between Brassica and other angiosperm lineages. J Mol Evol,1999,48:597-604.
123.Yano K, Takashi T, Nagamatsu S, Kojima M, Sakakibara H, Kitano H, Matsuoka M, Aya K. Efficacy of microarray profiling data combined with QTL mapping for the identification of a QTL gene controlling the initial growth rate in rice. Plant Cell Physiol,2012,53:729-739.
124.Zeng X, Zhu L, Chen Y, Qi L, Pu Y, Wen J, Yi B, Shen J, Ma C, Tu J, Fu T. Identification, fine mapping and characterisation of a dwarf mutant (bnaC.dwf) in Brassica napus. Theor Appl Genet, 2011, 122:421-428.
125.Zeng ZB. Precision mapping of quantitative trait loci. Genetics,1994,136: 1457-1468.
126.Zhao H, Shi L, Duan X, Xu F, Wang Y, Meng J. Mapping and validation of chromosome regions conferring a new boron-efficient locus in Brassica napus. Mol Breed, 2008, 22:495-506.
127.Zhao Z, Wu L, Nian F, Ding G, Shi T, Zhang D, Shi L, Xu F, Meng J. Dissecting quantitative trait loci for boron efficiency across multiple environments in Brassica napus. PLoS ONE,2012,7:e45215.