白菜与甘蓝型油菜—萝卜F染色体附加系回交BC1中萝卜F染色体的精准鉴定

doi:10.16420/j.issn.0513-353x.2021-0142

摘要/Abstract

摘要：

参考萝卜染色体定位序列标记连锁图谱中F染色体连锁群的SSR分子标记,利用21个已知萝卜染色体组成的甘蓝型油菜—萝卜附加系材料,确定了萝卜F染色体特异的2个SSR标记RM-59-293和Rs1SSR3694-220,应用两个标记对白菜‘559’与甘蓝型油菜—萝卜F染色体附加系FF回交的BC₁进行鉴定,筛选出10个含有萝卜F染色体的BC₁单株。利用萝卜基因组特异探针pURsN对该10个含有萝卜F染色体的BC₁单株的基因组进行荧光原位杂交,结果显示10个BC₁均得到1个杂交信号,含有1条萝卜F染色体。进一步对该10个BC₁单株进行主要农艺性状鉴定,筛选出形态接近白菜亲本‘559’的BC₁-15单株和形态偏向白菜类的BC₁-8、BC₁-16、BC₁-31和BC₁-35单株。

关键词: 白菜, 附加系, 萝卜F染色体, SSR分子标记, 荧光原位杂交

Abstract:

Referring to the SSR molecular markers of chromosome F linkage group in the chromosome-assigned sequence-tagged linkage map of radish,two SSR molecular markers RM-59-293 and Rs1SSR3694-220 which were specific for radish chromosome F were confirmed by means of the known radish-chromosome information in the genetic background of 21 materials. Both SSR markers were used to identify the alien radish chromosome F in the genetic background of BC₁ being derived from the backcross of Brassica rapa‘559’and rape-radish chromosome F addition line FF. As a result,10 BC₁ plants added with alien radish chromosome F were obtained. Furthermore,the genome of the selected 10 BC₁ plants with radish chromosome F were analyzed to detect the exogenous radish chromosome by fluorescence in situ hybridization technology using the radish genomic probe pURsN. The result showed that all of the 10 BC₁had one hybridization signal,which indicated that each of the tested BC₁ plants was added with an exogenous radish chromosome. The main agronomic traits of the 10 BC₁ plants were further identified. As a result,BC₁-15 with the morphology similar to Chinese cabbage parent‘559’and BC₁-8,BC₁-16,BC₁-31 and BC₁-35 with the morphology partial to Chinese cabbage were selected.

Key words: Brassica rapa, additional line, radish chromosome F, SSR molecular marker, fluorescent in situ hybridization

中图分类号:

丁云花, Budahn Holger, 赵泓, 赵岫云. 白菜与甘蓝型油菜—萝卜F染色体附加系回交BC₁中萝卜F染色体的精准鉴定[J]. 园艺学报, 2021, 48(7): 1295-1303.

DING Yunhua, Budahn Holger, ZHAO Hong, ZHAO Xiuyun. Accurate Identification of Radish Chromosome F in the Backcross Progeny of Brassica rapa and Rape-radish Chromosome F Addition Line[J]. Acta Horticulturae Sinica, 2021, 48(7): 1295-1303.

图/表 6

表1 SSR引物序列

Table 1 SSR primer sequences

名称 Name	引物序列（5′-3′） Primer sequence	片段/bp Product	基序 Motif	退火温度/℃ Annealing temperature
RM-59	F：TGTTAATCAGGTGGTTGTTACCC R：CCAAAGGAGAGAGCAAAGAGA	293	（AG）23	55
Rs1SSR3694	F：AGCGAGAAGATGAAGTAAAGTCTGA R：AAGTACCCAAAAGCATTAAAACCAT	220	（CT）6	55

图1 两对SSR引物RM-59（上）和Rs1SSR3694（下）对21个甘蓝型油菜—萝卜附加系相关标准材料的扩增结果 M：Marker;1、2：分别为萝卜亲本A24/2和A107/1;3 ~ 4：分别为甘蓝型油菜亲本A3/1和A246/1;5 ~ 12：分别为甘蓝型油菜—萝卜多体附加系14/52、14/15-66、14/32、14/53、14/35、14/37、14/45和14/15-61;13 ~ 21：分别为甘蓝型油菜—萝卜二体附加系AA、BB、CC、DD、EE、FF、GG、HH和II。

Fig. 1 Amplification results by two pairs of SSR-primers RM-59（above）and Rs1SSR3694（below）on the templates of 21 standard samples related to rape-radish addition lines M：Marker;1,2：Radish parents A24/2 and A107/1 respectively;3-4：Rape parents A3/1和A246/1 respectively;5-12：Multiple rape-radish chromosome additions 14/52,14/15-66,14/32,14/53,14/35,14/37,14/45 and 14/15-61 respectively;13-21：Disomic rape-radish chromosome addition lines AA,BB,CC,DD,EE,FF,GG,HH and II respectively.

图2 BC1中萝卜F染色体的SSR特异标记RM-59-293（上）和Rs1SSR3694-220（下）的鉴定结果 M：Marker;1：萝卜A107/1;2：甘蓝型油菜—萝卜F染色体附加系FF;3：白菜‘559’;4：FF与559杂交的F1;5 ~ 42：分别为38个BC1单株（1 ~ 38）。

Fig. 2 Identification results of radish chromosome F in the BC1 by SSR specific markers RM-59-293（above）and Rs1SSR3694-220（below） M：Marker;1：Radish A107/1;2：Rape-radish chromosome F addition line FF;3：Brassica rapa‘559’;4：F1 of FF and 559;5-42：38 BC1 individuals respectively（1-38）.

图3 用萝卜基因组特异探针pURsN检测甘蓝型油菜—萝卜F染色体附加系FF与白菜‘559’的BC1代的原位杂交结果

Fig. 3 Results tested by fluorescence in situ hybridization with radish specific probe pURsN on BC1 derived from rape-radish chromosome F additional line FF and Brassica rapa‘559’

表2 附加萝卜F染色体的BC1主要农艺性状调查

Table 2 Investigation on main agronomic traits of BC1 with additional radish F chromosome

编号 Code	叶形 Leaf shape	叶色 Leaf color	叶面 Blade surface	叶缘 Leaf margin
甘蓝型油菜—萝卜F染色体附加系FF Rape-radish chromosome F addition line FF	长椭圆Prolate ellipse	深绿Dark green	皱缩Crumpled	锯齿Jagged
白菜‘559’ Brassica rapa‘559’	宽椭圆Wide ellipse	绿Green	平滑Smooth	全缘Regular
BC₁-2	卵圆Oval ellipse	绿Green	平滑Smooth	锯齿Jagged
BC₁-8	长椭圆Prolate ellipse	浅绿Light green	平滑Smooth	锯齿Jagged
BC₁-15	倒卵圆Reverse oval ellipse	浅绿Light green	平滑Smooth	锯齿Jagged
BC₁-16	椭圆Ellipse	绿Green	平滑Smooth	锯齿Jagged
BC₁-21	椭圆Ellipse	绿Green	平滑Smooth	锯齿Jagged
BC₁-25	椭圆Ellipse	绿Green	皱缩Crumpled	锯齿Jagged
BC₁-28	宽椭圆Wide ellipse	绿Green	平滑Smooth	锯齿Jagged
BC₁-31	椭圆Ellipse	浅绿Light green	平滑Smooth	锯齿Jagged
BC₁-35	椭圆Ellipse	浅绿Light green	平滑Smooth	锯齿Jagged
BC₁-36	长椭圆Prolate ellipse	深绿Dark green	平滑Smooth	锯齿Jagged
编号 Code	叶裂刻 Leave lobe	叶尖形状 Leaf tip shape	叶面光泽 Foliar luster	叶翼 Blade base lobe
甘蓝型油菜—萝卜F染色体附加系FF Rape-radish chromosome F addition line FF	深裂Deep-lobe	钝尖Blunt	无Without	无Without
白菜‘559’ Brassica rapa‘559’	无裂刻No-lobe	圆Round	有With	有With
BC₁-2	无裂刻No-lobe	圆Round	无Without	无Without
BC₁-8	无裂刻No-lobe	圆Round	无Without	无Without
BC₁-15	无裂刻No-lobe	阔圆Broad round	有With	有With
BC₁-16	无裂刻No-lobe	钝尖Blunt	无Without	无Without
BC₁-21	无裂刻No-lobe	钝尖Blunt	无Without	无Without
BC₁-25	无裂刻No-lobe	阔圆Broad round	无Without	无Without
BC₁-28	无裂刻No-lobe	阔圆Broad round	无Without	无Without
BC₁-31	无裂刻No-lobe	钝尖Blunt	无Without	无Without
BC₁-35	无裂刻No-lobe	钝尖Blunt	无Without	有With
BC₁-36	无裂刻No-lobe	钝尖Blunt	无Without	无Without

表2 附加萝卜F染色体的BC1主要农艺性状调查

Table 2 Investigation on main agronomic traits of BC1 with additional radish F chromosome

编号 Code	叶形 Leaf shape	叶色 Leaf color	叶面 Blade surface	叶缘 Leaf margin
甘蓝型油菜—萝卜F染色体附加系FF Rape-radish chromosome F addition line FF	长椭圆Prolate ellipse	深绿Dark green	皱缩Crumpled	锯齿Jagged
白菜‘559’ Brassica rapa‘559’	宽椭圆Wide ellipse	绿Green	平滑Smooth	全缘Regular
BC₁-2	卵圆Oval ellipse	绿Green	平滑Smooth	锯齿Jagged
BC₁-8	长椭圆Prolate ellipse	浅绿Light green	平滑Smooth	锯齿Jagged
BC₁-15	倒卵圆Reverse oval ellipse	浅绿Light green	平滑Smooth	锯齿Jagged
BC₁-16	椭圆Ellipse	绿Green	平滑Smooth	锯齿Jagged
BC₁-21	椭圆Ellipse	绿Green	平滑Smooth	锯齿Jagged
BC₁-25	椭圆Ellipse	绿Green	皱缩Crumpled	锯齿Jagged
BC₁-28	宽椭圆Wide ellipse	绿Green	平滑Smooth	锯齿Jagged
BC₁-31	椭圆Ellipse	浅绿Light green	平滑Smooth	锯齿Jagged
BC₁-35	椭圆Ellipse	浅绿Light green	平滑Smooth	锯齿Jagged
BC₁-36	长椭圆Prolate ellipse	深绿Dark green	平滑Smooth	锯齿Jagged
编号 Code	叶裂刻 Leave lobe	叶尖形状 Leaf tip shape	叶面光泽 Foliar luster	叶翼 Blade base lobe
甘蓝型油菜—萝卜F染色体附加系FF Rape-radish chromosome F addition line FF	深裂Deep-lobe	钝尖Blunt	无Without	无Without
白菜‘559’ Brassica rapa‘559’	无裂刻No-lobe	圆Round	有With	有With
BC₁-2	无裂刻No-lobe	圆Round	无Without	无Without
BC₁-8	无裂刻No-lobe	圆Round	无Without	无Without
BC₁-15	无裂刻No-lobe	阔圆Broad round	有With	有With
BC₁-16	无裂刻No-lobe	钝尖Blunt	无Without	无Without
BC₁-21	无裂刻No-lobe	钝尖Blunt	无Without	无Without
BC₁-25	无裂刻No-lobe	阔圆Broad round	无Without	无Without
BC₁-28	无裂刻No-lobe	阔圆Broad round	无Without	无Without
BC₁-31	无裂刻No-lobe	钝尖Blunt	无Without	无Without
BC₁-35	无裂刻No-lobe	钝尖Blunt	无Without	有With
BC₁-36	无裂刻No-lobe	钝尖Blunt	无Without	无Without

图4 甘蓝型油菜—萝卜F染色体附加系FF、白菜‘559’及其附加萝卜F染色体的BC1植株

Fig. 4 Rape-radish chromosome F addition line FF,Brassica rapa‘559’and the BC1 plants with additional radish chromosome F

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白菜与甘蓝型油菜—萝卜F染色体附加系回交BC₁中萝卜F染色体的精准鉴定

Accurate Identification of Radish Chromosome F in the Backcross Progeny of Brassica rapa and Rape-radish Chromosome F Addition Line

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