Welcome to Acta Agronomica Sinica,

Acta Agronomica Sinica ›› 2022, Vol. 48 ›› Issue (3): 759-769.doi: 10.3724/SP.J.1006.2022.14025

• RESEARCH NOTES • Previous Articles     Next Articles

Development of linkage InDel markers of the white petal gene based on whole-genome re-sequencing data in Brassica napus L.

WANG Rui1,2(), CHEN Xue1,2, GUO Qing-Qing1,2, ZHOU Rong1,2, CHEN Lei1,2, LI Jia-Na1,2,*()   

  1. 1College of Agronomy and Biotechnology, Southwest University, Chongqing 400715, China
    2Chongqing Engineering Research Center for Rapeseed, Chongqing 400715, China
  • Received:2021-02-08 Accepted:2021-07-12 Online:2021-08-09 Published:2021-08-09
  • Contact: LI Jia-Na E-mail:Ruiwang71@163.com;ljn1950@swu.edu.cn
  • Supported by:
    Project of Intellectual Base for Discipline Innovation in Colleges and Universities (111 Program)(B12006)


InDel is widely distributed across the genome and occurs in a high density and large numbers in a genome. To date, the researches about linkage InDel markers of the white petal gene in B. napus L are very less yet. In this study, we constructed the F2 mapping population from the cross between DH Y05 (yellow petal) and DH W01 (white petal). Two bulks with 30 yellow petal lines and 30 white petal lines of F2 population were constructed by mixing an equal amount of DNA. Then two bulks and parents were performed 30× whole-genome re-sequencing. Darmor-bzh as the reference genome was aligned to sequence data from the two bulks and parents. QTL-seq and PoPoolation2 workflow were applied to identify the candidate region of the white petal gene. A major candidate region was identified on chromosome C03 (52-54 Mb) of Darmor-bzh. The insertion-deletion (InDel) sites can be visualized in candidate interval by Integrative Genomics Viewer (IGV). Based on these Indel variations, we used Vector and Blast to design InDel primers. Eight InDel markers closely linked to the white petal gene were screened by Polyacrylamide gel electrophoresis (PAGE). In summary, these results provide a basis for fine mapping white petal gene and InDel molecular marker located on functional genes as well as molecular marker assisted selection breeding.

Key words: Brassica napus L., re-sequencing, white petal genes, InDel markers

Fig. 1

∆(SNP-index) distribution on each chromosome Blue dot: ∆(SNP-index); Red line: sliding window average of ∆(SNP-index); Green lines: sliding window average of 95%-confidence interval upper/lower side; Orange line: sliding window average of 99%-confidence interval upper/lower side."

Fig. 2

White genes located on chromosome C03 (Chr. 13) using PoPoolation2"

Fig. 3

Visualization of InDel variation in candidate interval by IGV A: InDel-2; B: InDel-3; C: InDel-4; D: InDel-5; E: InDel-6; F: InDel-8; G: InDel-9; H: InDel-10."

Table 1

Primer information for InDels"

Primer name
Chr. C03 InDel 正向引物序列
Forward sequence (5°-3°)
Reverse sequence (5°-3°)

Fig. 4

Electrophoresis of parents and individuals of F2 population A: InDel-2; B: InDel-3; C: InDel-4; D: InDel-5; E: InDel-6; F: InDel-8; G: InDel-9; H: InDel-10; M: 20 bp ladder. 1: parents with yellow petal; 2: parents with white petal; 3-13: 11 yellow petal plants of F2; 14-24: 11 white petal plants of F2."

[1] 王汉中. 以新需求为导向的油菜产业发展战略. 中国油料作物学报, 2018, 40:613-617.
Wang H Z. New-demand oriented oilseed rape industry developing strategy. Chin J Oil Crop Sci, 2018, 40:613-617 (in Chinese with English abstract).
[2] 阴长发, 官春云. 油菜花色研究进展. 作物研究, 2013, 27:403-408.
Yin C F, Guan C Y. A review of rapeseed flower color. Crop Res, 2013, 27:403-408 (in Chinese with English abstract).
[3] 刘后利. 油菜的遗传和育种. 上海: 上海科学技术出版社, 1985. pp 76-77.
Liu H L. Heredity and Breeding of Rape. Shanghai: Shanghai Scientific and Technical Publishers, 1985. pp 76-77(in Chinese)
[4] Chen B, Heneen W, Jonsson R. Brassica napus L Brassica napus L. Plant Breed, 1988, 100:147-149.
doi: 10.1111/pbr.1988.100.issue-2
[5] 戚存扣, 傅寿仲. 甘蓝型油菜白花性状的遗传. 中国油料作物学报, 1992, 1(3):60-62.
Qi C K, Fu S Z. Genetic studies of white petals in Brassica napus L. Chin J Oil Crop Sci, 1992, 1(3):60-62 (in Chinese with English abstract).
[6] Pearson O H. Brassica oleracea L Brassica oleracea L. Am Naturalist, 1929, 63:561-565.
doi: 10.1086/280291
[7] 张豹. 甘蓝型油菜导入系构建、重要农艺性状QTL分析和白花基因克隆. 华中农业大学博士学位论文, 湖北武汉, 2015.
Zhang B. Development of Chromosome Segment Substitution Lines for QTL Analysis of Important Agronomic Traits and Cloning the White-flowered Gene in Brassica napus L. PhD Dissertation of Huazhong Agricultural University, Wuhan, Hubei, China, 2015 (in Chinese with English abstract).
[8] 张洁夫, 浦惠明, 戚存扣, 傅寿仲. 甘蓝型油菜花色性状的遗传研究. 中国油料作物学报, 2000, 22(3):1-4.
Zhang J F, Pu H M, Qi C K, Fu S Z. Inheritance of flower color character in oilseed rape Brassica napus L. Chin J Oil Crop Sci, 2000, 22(3):1-4 (in Chinese with English abstract).
[9] 王翊, 景尚友, 吴刚, 任丽杰. 甘蓝型油菜白花性状在杂交油菜育种中的应用. 黑龙江农业科学, 2003, (6):13-14.
Wang Y, Jing S Y, Wu G, Ren L J. Application of the character of white flower of Brassica napus L. in hybrid breeding. Heilongjiang Agric Sci, 2003, (6):13-14 (in Chinese with English abstract).
[10] 文雁成, 张书芬, 王建平, 朱家成, 赵磊. 甘蓝型油菜白花性状的遗传学研究和白花胞质雄性不育系的选育. 中国农学通报, 2010, 26(1):95-97.
Wen Y C, Zhang S F, Wang J P, Zhu J C, Zhao L. Genetic studies of white Petals and selection of cytoplasmic male sterile line with white petals in Brassica napus L. Chin Agric Sci Bull, 2010, 26(1):95-97 (in Chinese with English abstract).
[11] 黄镇, 许婷, 班元元, 刘欢, 范胜栩, 杨丽, 徐爱遐. 甘蓝型油菜白花性状的遗传及AFLP标记. 华北农学报, 2012, 27(1):98-101.
Huang Z, Xu T, Ban Y Y, Liu H, Fan S X, Yang L, Xu A X. Genetic studies of white petals and AFLP markers linked to white petal gene in Brassica napus L. Acta Agric Boreali-Sin, 2012, 27(1):98-101 (in Chinese with English abstract).
[12] 邓昌蓉, 赵志刚, 余青兰. 人工合成甘蓝型油菜花色变异后代的遗传研究. 北方园艺, 2014, (18):14-17.
Deng C R, Zhao Z G, Yu Q L. The genetic studies of the flower color variation’s offsprings in artificial synthesis of Brassica napus. Northern Hortic, 2014, (18):14-17 (in Chinese with English abstract).
[13] 董育红, 田建华, 李殿荣, 郭蔼光, 孔建, 赵小萍. 甘蓝型油菜白花基因的RAPD标记. 西北农林科技大学学报, 2005, 33(10):57-61.
Dong Y H, Tian J H, Li D R, Guo A G, Kong J, Zhao X P. RAPD markers linked to white-petal gene in Brassica napus L. J Northwest A&F Univ, 2005, 33(10):57-61 (in Chinese with English abstract).
[14] 黄萌, 张建栋, 陈培峰, 宋英, 孙华. 甘蓝型油菜白花性状的遗传规律. 江苏农业科学, 2017, 45(20):83-84.
Huang M, Zhang J D, Chen P F, Song Y, Sun H. Genetic law of white flower traits in Brassica napus L. Jiangsu Agric Sci, 2017, 45(20):83-84 (in Chinese with English abstract).
[15] 田露申, 牛应泽, 余青青, 郭世星, 柳丽. 甘蓝型油菜白花性状的主基因+多基因遗传分析. 中国农业科学, 2009, 42:3987-3995.
Tian L S, Niu Y Z, Yu Q Q, Guo S X, Liu L. Genetic analysis of white flower color with mixed model of major gene plus polygene in Brassica napus L. Sci Agric Sin, 2009, 42:3987-3995 (in Chinese with English abstract).
[16] 刘雪平, 涂金星, 陈宝元, 傅廷栋. 人工合成甘蓝型油菜中花色与芥酸含量的遗传连锁分析. 遗传学报, 2004, 31:357-362.
Liu X P, Tu J X, Chen B Y, Fu T D. Identification of the linkage relationship between the flower colour and the content of erucic acid in the resynthesized Brassica napus L. Acta Genet Sin, 2004, 31:357-362 (in English with Chinese abstract).
[17] Han F Q, Yang C, Fang Z Y, Yang L M, Zhang M, Lyu H H, Liu Y M, Li Z S, Liu B, Yu H L, Liu X P, Zhang Y Y. cpc-1) in Brassica oleracea cpc-1) in Brassica oleracea. Mol Breed, 2015, 35:160.
doi: 10.1007/s11032-015-0354-x
[18] Huang Z, Ban Y Y, Bao R, Zhang X X, Xu A X, Ding J. Brassica napus L Brassica napus L. New Zealand J Crop Hortic Sci, 2014, 111:117.
[19] Zhang X X, Li R H, Niu S L, Chen L, Gao J, Wen J, Yi B, Ma C Z, Tu J X, Fu T D, Shen J X. Brassica juncea white-flowered mutant Bjpc2 using the whole-genome resequencing Brassica juncea white-flowered mutant Bjpc2 using the whole-genome resequencing. Mol Genet Genomics, 2017, 293:359-370.
doi: 10.1007/s00438-017-1390-5
[20] Xiao S, Xu J, Li Y, Zhang L, Shi S, Shi S, Wu J, Liu K. Brassica napus using a genome-walking technique Brassica napus using a genome-walking technique. Genome, 2007, 50:611-618.
doi: 10.1139/G07-044
[21] 陈雪, 王瑞, 井付钰, 张胜森, 贾乐东, 段谋正, 吴宇. 基于二代测序的甘蓝型油菜白花基因候选区间定位及连锁标记验证. 中国农业科学, 2020, 53:1108-1117.
Chen X, Wang R, Jing F Y, Zhang S S, Jia L D, Duan M Z, Wu Y. Location and linkage markers for candidate interval of the white petal gene in Brassica napus L. by next generation sequencing. Sci Agric Sin, 2020, 53:1108-1117 (in Chinese with English abstract).
[22] Zhang B, Liu C, Wang Y, Yao X, Wang F, Wu J, King G J, Liu K. Brassica species Brassica species. New Phytol, 2015, 206:1513-1526.
doi: 10.1111/nph.13335 pmid: 25690717
[23] Yao Y M, Li K X, Liu H D, Duncan R W, Guo S M, Xiao L, Du D Z. Bnpc 1) in spring Brassica napus L. to a 151-kb region Bnpc 1) in spring Brassica napus L. to a 151-kb region. Euphytica, 2017, 213:165.
doi: 10.1007/s10681-017-1959-4
[24] 丁戈, 陈伦林, 邹小云, 李书宇, 熊洁, 邹晓芬, 宋来强. 甘蓝型油菜桔黄花色基因的QTL-seq遗传分析及InDel分子标记开发. 分子植物育种, 2019, 17:3983-3992.
Ding G, Chen L L, Zou X Y, Li S Y, Xiong J, Zou X F, Song L Q. QTL-seq genetic analysis and InDel marker development of orange petel color gene in Brassica napus. Mol Plant Breed, 2019, 17:3983-3992 (in Chinese with English abstract).
[25] Mithra S V A, Kar M K, Mohapatra T, Robin S, Sarla N, Seshashayee M, Singh K, Singh N K, Sharma R P. DBT propelled national effort in creating mutant resource for functional genomics in rice. Curr Sci, 2016, 110:543-548.
doi: 10.18520/cs/v110/i4/543-548
[26] Wei F J, Droc G, Guiderdoni E, Hsing Y I C. International consortium of rice mutagenesis: Resources and beyond. Rice, 2013, 6:39.
doi: 10.1186/1939-8433-6-39
[27] Tsuda M, Kaga A, Anai T, Shimizu T, Sayamat, Takagi K, Machita K, Watanabe S, Nishimura M, Yamada N, Mori S, Sasaki H, Kanamori H, Katayose Y, Ishimoto M. Construction of a high-density mutant library in soybean and development of a mutant retrieval method using amplicon sequencing. BMC Genomics, 2015, 16:1014.
doi: 10.1186/s12864-015-2079-y pmid: 26610706
[28] Just D, Garcia V, Fernandez L, Bres C, Mauxion J P, Petit J, Jorly J, Assali J, Bournonville C, Ferrand C, Baldet P, Lemaire- Chamley M, Mori K, Okabe Y, Ariizumi T, Asamizu E, Ezura H, Rothan C. Micro-Tom mutants for functional analysis of target genes and discovery of new alleles in tomato. Plant Biotechnol J, 2013, 30:225-231.
[29] Lin T, Wang S H, Zhong Y, Gao D L, Cui Q Z, Chen H M, Zhang Z H, Shen H L, Weng Y Q, Huang S W. A truncated F-box protein confers the dwarfism in cucumber. J Genet Genomics, 2016, 43:223-226.
doi: 10.1016/j.jgg.2016.01.007
[30] Lun Y Y, Wang X, Zhang C Z, Yang L, Gao D L, Chen H M, Huang S W. A CsYcf54 variant conferring light green coloration in cucumber. Euphytica, 2016, 208:509-517.
doi: 10.1007/s10681-015-1592-z
[31] Zhou Q, Wang S H, Hu B W, Chen H M, Zhang Z H, Huang S W. An accumulation and replication of chloroplasts 5 gene mutation confers light green peel in cucumber. J Integr Plant Biol, 2015, 57:936-942.
doi: 10.1111/jipb.12355
[32] Takagi H, Abe A, Yoshid A K, Kosugi S, Natsume S, Mitsuoka C, Uemura A, Utsushi H, Tamiru M, Takumo S, Innan H, Cano L M, Kamoun S, Terauchi R. QTL-seq: rapid mapping of quantitative trait loci in rice by whole genome resequencing of DNA from two bulked populations. Plant J, 2013, 74:174-183.
doi: 10.1111/tpj.2013.74.issue-1
[33] Robert K, Ram Vinay P, Christian S. PoPoolation2: Identifying differentiation between populations using sequencing of pooled DNA samples (Pool-Seq). Bioinformatics, 2011, 27:3435-3436.
doi: 10.1093/bioinformatics/btr589 pmid: 22025480
[34] Hua Y P, Zhang D D, Zhou T, He M L, Ding G D, Shi L, Xu F S. Transcriptomics-assisted quantitative trait locus fine mapping for the rapid identification of a nodulin 26-like intrinsic protein gene regulating boron efficiency in allotetraploid rapeseed. Plant Cell Environ, 2016, 39:1601-1618.
doi: 10.1111/pce.v39.7
[35] 淡亚彬. 甘蓝型油菜桔红花色基因和心叶紫色基因的初步定位. 青海大学硕士学位论文, 青海西宁, 2016.
Dan Y B. Primary Mapping of the Orange Flower Gene and Central Leaf Color Gene in Brassica napus L. MS Thesis of Qinghai University, Xining, Qinghai, China, 2016 (in Chinese with English abstract)
[36] 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, 21:447-453.
Pan C H, Wang Z B, Ma Y Y, Yin Y J, Zhang Y F, Zuo S M, Chen Z X, Pan X B. InDel and SNP markers and their application in map-based cloning of rice genes. Chin J Rice Sci, 2007, 21:447-453 (in Chinese with English abstract).
[37] 胡坤. 玉米与大刍草InDel标记的开发及遗传连锁图谱的构建. 四川农业大学硕士学位论文, 四川雅安, 2014.
Hu K. The Development of Maize and Teosinte InDel Markers and Genetic Linkage Map Construction. MS Thesis of Sichuan Agricultural University, Ya’an, Sichuan, China, 2014 (in Chinese with English abstract).
[38] 吴迷, 汪念, 沈超, 黄聪, 温天旺, 林忠旭. 基于重测序的陆地棉InDel标记开发与评价. 作物学报, 2019, 45:196-203.
doi: 10.3724/SP.J.1006.2019.84100
Wu M, Wang N, Shen C, Huang C, Wen T W, Lin Z X. Development and evaluation of InDel markers in cotton based on whole-genome re-sequencing data. Acta Agron Sin, 2019, 45:196-203 (in Chinese with English abstract).
[39] 徐婷婷, 汪巧玲, 邹淑琼, 狄佳春, 杨欣, 朱银, 赵涵, 颜伟. 基于高通量测序的大麦InDel标记开发及应用. 作物学报, 2020, 46:1340-1355.
doi: 10.3724/SP.J.1006.2020.91076
Xu T T, Wang Q L, Zou S Q, Di J C, Yang X, Zhu Y, Zhao H, Yan W. Development and application of InDel markers based on high throughput sequencing in barley. Acta Agron Sin, 2020, 46:1340-1355 (in Chinese with English abstract).
[40] 周新桐, 郭青青, 陈雪, 李加纳, 王瑞. GBS高密度遗传连锁图谱定位甘蓝型油菜粉色花性状. 作物学报, 2021, 47:587-598.
doi: 10.3724/SP.J.1006.2021.04115
Zhou X T, Guo Q Q, Chen X, Li J N, Wang R. Construction of a high-density genetic map using genotyping by sequencing (GBS) for quantitative trait loci (QTL) analysis of pink petal trait in Brassica napus L. Acta Agron Sin, 2021, 47:587-598 (in Chinese with English abstract).
[1] ZHOU Xin-Tong, GUO Qing-Qing, CHEN Xue, LI Jia-Na, WANG Rui. Construction of a high-density genetic map using genotyping by sequencing (GBS) for quantitative trait loci (QTL) analysis of pink petal trait in Brassica napus L. [J]. Acta Agronomica Sinica, 2021, 47(4): 587-598.
[2] LI Shu-Yu, HUANG Yang, XIONG Jie, DING Ge, CHEN Lun-Lin, SONG Lai-Qiang. QTL mapping and candidate genes screening of earliness traits in Brassica napus L. [J]. Acta Agronomica Sinica, 2021, 47(4): 626-637.
[3] MENG Jiang-Yu, LIANG Guang-Wei, HE Ya-Jun, QIAN Wei. QTL mapping of salt and drought tolerance related traits in Brassica napus L. [J]. Acta Agronomica Sinica, 2021, 47(3): 462-471.
[4] WANG Rui-Li, WANG Liu-Yan, LEI Wei, WU Jia-Yi, SHI Hong-Song, LI Chen-Yang, TANG Zhang-Lin, LI Jia-Na, ZHOU Qing-Yuan, CUI Cui. Screening candidate genes related to aluminum toxicity stress at germination stage via RNA-seq and QTL mapping in Brassica napus L. [J]. Acta Agronomica Sinica, 2021, 47(12): 2407-2422.
[5] GUO Qing-Qing, ZHOU Rong, CHEN Xue, CHEN Lei, LI Jia-Na, WANG Rui. Location and InDel markers for candidate interval of the orange petal gene in Brassica napus L. by next generation sequencing [J]. Acta Agronomica Sinica, 2021, 47(11): 2163-2172.
[6] LEI Wei, WANG Rui-Li, WANG Liu-Yan, YUAN Fang, MENG Li-Jiao, XING Ming-Li, XU Lu, TANG Zhang-Lin, LI Jia-Na, CUI Cui, ZHOU Qing-Yuan. Genome-wide association study of seed density and its related traits in Brassica napus L. [J]. Acta Agronomica Sinica, 2021, 47(11): 2099-2110.
[7] XU Ting-Ting, WANG Qiao-Ling, ZOU Shu-Qiong, DI Jia-Chun, YANG Xin, ZHU Yin, ZHAO Han, YAN Wei. Development and application of InDel markers based on high throughput sequencing in barley [J]. Acta Agronomica Sinica, 2020, 46(9): 1340-1350.
[8] JIANG Shu-Kun,WANG Li-Zhi,YANG Xian-Li,LI Bo,MU Wei-Jie,DONG Shi-Chen,CHE Wei-Cai,LI Zhong-Jie,CHI Li-Yong,LI Ming-Xian,ZHANG Xi-Juan,JIANG Hui,LI Rui,ZHAO Qian,LI Wen-Hua. Detection of QTLs controlling cold tolerance at bud bursting stage by using a high-density SNP linkage map in japonica rice [J]. Acta Agronomica Sinica, 2020, 46(8): 1174-1184.
[9] WANG Rui-Li,WANG Liu-Yan,YE Sang,Gao Huan-Huan,LEI Wei,WU Jia-Yi,YUAN Fang,MENG Li-Jiao,TANG Zhang-Lin,LI Jia-Na,ZHOU Qing-Yuan,CUI Cui. QTL mapping of seed germination-related traits in Brassica napus L. under aluminum toxicity stress [J]. Acta Agronomica Sinica, 2020, 46(6): 832-843.
[10] LYU Wei-Sheng, XIAO Fu-Liang, ZHANG Shao-Wen, ZHENG Wei, HUANG Tian-Bao, XIAO Xiao-Jun, LI Ya-Zhen, WU Yan, HAN De-Peng, XIAO Guo-Bin, ZHANG Xue-Kun. Effects of sowing and fertilizing methods on yield and fertilizer use efficiency in red-soil dryland rapeseed (Brassica napus L.) [J]. Acta Agronomica Sinica, 2020, 46(11): 1790-1800.
[11] HU Mao-Long, CHENG Li, GUO Yue, LONG Wei-Hua, GAO Jian-Qin, PU Hui-Ming, ZHANG Jie-Fu, CHEN Song. Development and application of the marker for imidazolinone-resistant gene in Brassica napus [J]. Acta Agronomica Sinica, 2020, 46(10): 1639-1646.
[12] Cun-Min QU,Guo-Qiang MA,Mei-Chen ZHU,Xiao-Hu HUANG,Le-Dong JIA,Shu-Xian WANG,Hui-Yan ZHAO,Xin-Fu XU,Kun LU,Jia-Na LI,Rui WANG. Genome-wide association of roots, hypocotyls and fresh weight at germination stage under as stress in Brassica napus L. [J]. Acta Agronomica Sinica, 2019, 45(2): 175-187.
[13] Xiao-Ding MA,Jiang-Hong TANG,Jia-Ni ZHANG,Di CUI,Hui LI,Mao-Mao LI,Long-Zhi HAN. Development of molecular markers polymorphic between Dongxiang wild rice and Geng rice cultivar ‘Nipponbare’ [J]. Acta Agronomica Sinica, 2019, 45(2): 316-321.
[14] Yang-Yang LI,Rong-Rong JING,Rong-Rong LYU,Peng-Cheng SHI,Xin LI,Qin WANG,Dan WU,Qing-Yuan ZHOU,Jia-Na LI,Zhang-Lin TANG. Genome-wide association analysis and candidate genes prediction of waterlogging-responding traits in Brassica napus L. [J]. Acta Agronomica Sinica, 2019, 45(12): 1806-1821.
[15] Tao FENG,Chun-Yun GUAN. Cloning and Characterization of Brassinazole-resistant (BnaBZR1 and BnaBES1) CDS from Brassica napus L. [J]. Acta Agronomica Sinica, 2018, 44(12): 1793-1801.
Full text



No Suggested Reading articles found!