VNTR特异性引物在甜菜育性鉴定中的应用

doi:10.7668/hbnxb.20192531

摘要/Abstract

摘要：

为了摸清甜菜选育进程中部分不育系和保持系材料的细胞质类型,为下一步的选育工作提供指导方向。利用分子标记引物TR1对国内总共30份甜菜试验材料开展细胞质类型的分子水平鉴定,并结合田间花期育性性状调查进行比对。结果表明,成型稳定的保持系960767和不育系N9848经过TR1引物鉴定后的PCR电泳条带分别为750,500 bp,其胞质类型分别为N1和S型,其他试验材料检测到750 bp的材料为T152、T328、T334、T306、T360,推断其为N1型细胞质;检测到500 bp条带的材料为T766、T154、N122、I-1、B-3、Z-1、41419、10320、B-3-23、S865、S301、S151、S327、S333、S115、S305、S153、S359、S137、S163和S313,推断其为S型细胞质;检测到500~750 bp条带的材料为T302、T116。由于T302在田间调查中其对S301的不育性也具有保持能力,因此将T302和T116分为N型细胞质的另一种类型N2。同时检测到500 bp和750 bp的材料为T302、I-1和B-3的部分植株,为同时包括正常和欧文细胞质的材料,属于特质性材料。田间花粉育性调查结果T766、T154、N122、I-1、B-3、Z-1、41419、10320、B-3-23均为可育型,但鉴定其胞质类型属于S型细胞质。经分子鉴定和田间育性性状调查对比,大部分材料分子鉴定结果与育性性状调查结果一致。部分材料存在分子鉴定与育性性状结果不一致,有待于进一步调整选育方向。

关键词: 甜菜, 分子标记, 育性鉴定, TR1引物

Abstract:

In order to find out the cytoplasm types of some sterile line and maintainer line materials in the process of sugar beet breeding, it provides a guiding direction for the next breeding work. Molecular marker primer TR1 was used to identify the molecular level of 30 test materials cytoplasm types, and to compare the fertility traits in the field. The results showed that the PCR electrophoresis bands of maintainer line 960767 and sterile line N9848 were 750,500 bp.We named these cytoplasm types as N1 and S,respectively.And 750 bp bands were detected by T152,T328,T334,T306,T360,as N1 cytoplasm.The 500 bp bands detected were T766,T154,N122,I-1,B-3,Z-1,41419,10320,B-3-23,S865,S301,S151,S327,S333,S115,S305,S153,S359,S137,S163,and S313,inferred to be S type cytoplasm.The materials for the bands detected between 500 bp and 750 bp were T302,and T116.Because T302 also had the ability to maintain the sterility of S301 in field investigations,T302 and T116 were classified into another type,N2 cytoplasm type.Some plants with 500, 750 bp of materials were T302, I-1 and B-3, which included both normal and Owen cytoplasm were characteristic materials. The results of field pollen fertility survey T766, T154, N122, I-1, B-3, Z-1, 414191, 10320 and B-3-23 were all fertile, but their cytoplasm type was identified to belong to S type cytoplasm. According to molecular identification and field fertility trait survey, the results of most materials were consistent with the results of fertility trait survey. Some materials had inconsistent molecular identification and fertility trait results, requiring further adjustment of the selection direction.

Key words: Sugar beet, Molecular markers, Fertility identification, TR1 primer

张辉, 王良, 付增娟, 李晓东, 赵尚敏, 鄂圆圆, 郑文哲, 张自强, 张必周, 张惠忠. VNTR特异性引物在甜菜育性鉴定中的应用[J]. 华北农学报, 2022, 37(2): 35-41. doi: 10.7668/hbnxb.20192531.

ZHANG Hui, WANG Liang, FU Zengjuan, LI Xiaodong, ZHAO Shangmin, E Yuanyuan, ZHENG Wenzhe, ZHANG Ziqiang, ZHANG Bizhou, ZHANG Huizhong. Application of VNTR-specific Primers in Fertility Identification of Sugar Beet[J]. Acta Agriculturae Boreali-Sinica, 2022, 37(2): 35-41. doi: 10.7668/hbnxb.20192531.

http://www.hbnxb.net/CN/Y2022/V37/I2/35

图/表 8

表1 试验材料名称

Tab.1 Test materials names

序号 Number	材料名称 Material name	序号 Number	材料名称 Material name
1	960767	16	N9849
2	T766	17	S865
3	T302	18	S301
4	T152	19	S151
5	T328	20	S327
6	T334	21	S333
7	T116	22	S115
8	T306	23	S305
9	T154	24	S153
10	T360	25	S359
11	N122	26	S137
12	I-1	27	S163
13	B-3	28	S313
14	Z-1	29	10320
15	41419	30	B-3-23

图1 1%琼脂糖凝胶DNA检测
1—28.除去960767和N9849的其他试验材料的电泳检测结果。

Fig.1 1% agarose gel DNA detection
1—28.The electrophoresis detection results of the other test materials excluding 960767 and N9849.

图2 保持系960767和不育系N9849经TR1引物PCR扩增后的检测结果
1—8.保持系材料960767不同植株的TR1引物检测结果;9—17.不育系材料N9849不同植株的TR1引物检测结果。

Fig.2 Results of maintainer 960767 and sterile N9849 amplified by the TR1 primer PCR
1—8.The TR1 primer test results of different plants of maintainer material 960767;9—17.The TR1 primer test results of different plants of sterile line material N9849.

图3 部分材料经TR1引物PCR扩增后的检测结果

Fig.3 Detection results of some materials amplified by the TR1 primer PCR
1.T334;2.S333;4,5.S115;3,6.T116;8,10.T302;7,9.S301;11,12.960767;13—15.I-1;16—18.B-3;19—21.Z-1;22—24.41419;25—26.10320;27—28.B-3-23。

图4 保持系T302和不育系S301经TR1引物PCR扩增后的检测结果
1—5.不育型材料S301不同植株样品检测;6—10.不育型材料S115不同植株样品检测;11—15.N2型细胞质材料T302不同植株样品检测;16—20.N2型细胞质材料T116不同植株样品检测。

Fig.4 Detection results of maintainer material T302 and sterile material S301 amplified with the TR1 primer PCR
1—5.Detection of different plants of sterility material S301;6—10.Detection of different plants of sterile material S115;11—15.Detection of different plants of N2 cytoplasmic materials T302;16—20.Detection of different plants of N2 cytoplasmic materials T116.

图5 部分可育材料经TR1引物PCR扩增后的检测结果
1—6.960767不同植株检测;7—13.T152不同植株检测;14—20.T328不同植株检测;21—27.T334不同植株检测;28—31.T306不同植株检测;32—35.T360不同植株检测;36—40.T302不同植株检测;41—44.T116不同植株检测;45—48.T766不同植株检测;49—54.N122不同植株检测。

Fig.5 Detection results of some fertile materials amplified by the TR1 primer PCR
1—6.Detection of different plants of 960767;7—13.Detection of different plants T152;14—20.Detection of different plants T328;21—27.Detection of different plants T334;28—31.Detection of different plants T306;32—35.Detection of different plants T360;36—40.Detection of different plants T302;41—44.Detection of different plants T116;45—48.Detection of different plants T766;49—54.Detection of different plants N122.

图6 部分不育型材料经TR1引物PCR扩增后的检测结果
1—2.S865不同植株检测;3—4.S301不同植株检测;5—6.S151不同植株检测;7—8.S327不同植株检测;9—10.S333不同植株检测;11—12.S115不同植株检测;13—14.S305不同植株检测;15—16.S315不同植株检测;17—18.S359不同植株检测;19—20.S137不同植株检测;21—22.S163不同植株检测;23—24.S313不同植株检测。

Fig.6 Test results of some sterile materials by TR1 primer PCR
1—2.Detection of different plants of S865;3—4.Detection of different plants of S301;5—6.Detection of different plants of S151;7—8.Detection of different plants of S327;9—10.Detection of different plants of S333;11—12.Detection of different plants of S115;13—14.Detection of different plants of S305;15—16.Detection of different plants of S315;17—18.Detection of different plants of S359;19—20.Detection of different plants of S137;21—22.Detection of different plants of S163;23—24.Detection of different plants of S313.

表2 不同试验材料经TR1引物扩增鉴定及田间育性调查

Tab.2 Identification of amplification and field breeding investigation by TR1 primers of different test materials

材料名称 Material name	条带位置/bp Band position	育性性状 Fertility character	类型 Type	材料名称 Material name	条带位置/bp Band position	育性性状 Fertility character	类型 Type
960767	750	可育型	N1	N9849	500	全不育型	S
T766	500	可育型	S	S865	500	半不育二型	S
T302	500—750	可育型	N2	S301	500	全不育型	S
T152	750	可育型	N1	S151	500	全不育型	S
T328	750	可育型	N1	S327	500	全不育型	S
T334	750	可育型	N1	S333	500	全不育型	S
T116	500—750	可育型	N2	S115	500	半不育二型	S
T306	750	半可育型	N1	S305	500	半不育一型	S
T154	500	半可育型	S	S153	500	半不育二型	S
T360	750/500	可育型	N1	S359	500	全不育型	S
N122	500	可育型	S	S137	500	半不育二型	S
I-1	500	可育型	S	S163	500	半不育二型	S
B-3	500	可育型	S	S313	500	全不育型	S
Z-1	500	可育型	S	10320	500	可育型	S
41419	500	可育型	S	B-3-23	500	可育型	S

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