关中地区紫花苜蓿品种优势性比较研究
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摘要
我国是蛋白质饲料需求最多而缺口最大的国家,蛋白质饲料的严重不足将成为制约我国畜牧业发展的关键因素。豆科牧草紫花苜蓿(Medicago sativa L.)因兼具抗逆性强、产量高、品质好、利用方式多、适口性佳等优点,被公认为是世界上最大的植物蛋白库,大力培植“苜蓿型奶牛业”,是保障原料奶质量安全的唯一正确道路。陕西关中作为我国最早引种苜蓿的地区,在长期种植过程中存在品种混乱、遗传背景不清等问题,影响苜蓿的产业化发展。为此,本研究对关中地区22个国内外紫花苜蓿品种的亲缘关系进行了分子遗传标记分析和鉴定,理清其亲缘关系,同时在大田条件下,连续多年对各品种的农艺学性状、产量性状及生物学特性观察分析的基础上,筛选出具有较强杂种优势的7个品种,继续对其开花习性进行了观察,为种子生产和杂交做基础工作。最后,以金皇后为材料,对盐胁迫下紫花苜蓿抗氧系统的调节机制其进行了研究。
本研究取得的主要结论有:
1. 22个苜蓿品种的遗传多样性及亲缘关系分析
RAPD标记聚类关系结果表明,22个紫花苜蓿品种可以聚为三大类:引自美国的阿尔岗金和引自荷兰的三得利聚为第一大类,维多利亚、全能、皇后、金皇后、艾菲尼特这5个引自美国的品种聚为第二大类,我国的关中苜蓿、固原紫花、中苜一号和其余品种聚为第三大类。其中,中苜一号和固原紫花苜蓿属于同一亚类,二者之间亲缘关系相对较近,与关中苜蓿之间关系相对较远;加拿大品种放牧者独列一类,与其它紫花苜蓿品种亲缘关系均相对较远,说明用它与其它21个苜蓿品种杂交,均有可能产生明显的杂种优势。
2. 22个苜蓿品种的生产及品质特性比较
(1)不同苜蓿品种的生育时期具有一定的差异,返青期差异最明显,最早和最晚相差达15天,但随着生长的延长其差异间隔天数在缩短,到了开花期,品种间差异几乎不明显;
(2)播种当年头茬苜蓿株高、产量均较低,2茬、3茬较高。株高分析初步可以判断,品种维多利亚、全能、三得利、德福、苜蓿54、放牧者具有较好的生长潜势,而品种关中苜蓿、WL252HQ、固原紫花则相对表现较差;
(3)22个苜蓿品种连续三茬的生长速度差异显著(P<0.05),品种维多利亚、全能、三得利、德福、苜蓿54、放牧者较有优势。品种三得利和品种维多利亚的生长强度最高,关中苜蓿生长强度最低,与品种三得利和品种维多利亚相差近一倍;品种维多利亚、放牧者、WL324、苜蓿54、WH323ML头茬再生速度最快,其中品种维多利亚和品种放牧者达0.8cm/d以上,表现最差的仍为固原紫花和关中苜蓿,其头茬再生速度只有0.49 cm/d左右;
(4)方差分析结果表明,22个苜蓿品种间的鲜干草产量差异均达到极显著水平(P<0.01)。品种维多利亚的鲜草产量除了与全能、皇后、牧歌、阿尔冈金、中苜一号、WL324、苜蓿54、放牧者之间差异不显著外,均显著高出其他品种(P<0.05);
(5)品种WL324(12)单位面积的蛋白质产出量最高,达到3216.0 kg/hm2,比最低的关中苜蓿高出115.0%,除了与品种维多利亚、苜蓿54、放牧者差异不显著外,与其他品种差异均显著;
(6)品种间草株比具有极显著差异(P<0.01)。中苜一号、维多利亚、WL323HQ、WL232HQ、德福草株比最高,都在0.75以上,直立性最好;品种阿尔冈金、WH323ML、德宝最低,只有0.55-0.59。
综上,维多利亚、全能、三得利、爱非利特和放牧者5个优势品种和两个地方品种关中苜蓿、固原紫花被筛选出来并做为下一步杂交和种子生产习性研究的材料。
3. 7个紫花苜蓿品种的种子生产性能评价
(1)关中地区紫花苜蓿一般在7:00~19:00小花开放,9:00~15:00为龙骨瓣开放高峰期,是昆虫活动的高峰期,也是进行苜蓿杂交的最佳时期;
(2) 7个紫花苜蓿品种单个花序小花数、单个花序结荚数和单个荚内种子数相关性分析结果显示各指标之间的相关性因品种而异;
(3)关中苜蓿和固原紫花2个紫花苜蓿品种的单个花序小花数、单个花序结荚数和单个荚内种子数均不及国外引进的三得利、爱非尼特、放牧者、全能和维多利亚5个紫花苜蓿品种。
4.盐胁迫下紫花苜蓿抗氧化系统的调节机制
为研究盐胁迫下紫花苜蓿对逆境生理生化的调节机制,以紫花苜蓿幼苗为研究材料,采用不同NaCl浓度(0、40、80、120和160 mmol/L)进行盐胁迫处理,对其幼苗叶片和根部的超氧阴离子自由基( O_2~(·-) ),膜脂过氧化产物丙二醛(MDA),以及抗氧化系统酶中的过氧化物酶(POD),超氧化物歧化酶(SOD),过氧化氢酶(CAT)和非酶抗氧化剂Vc和总抗氧化能力(T-AOC)等指标进行了测定。研究结果表明,随着NaCl盐浓度增加,超氧阴离子自由基和MDA积累增加,抗氧化物酶POD、SOD和CAT活性也不断增加,同时非酶抗氧化剂Vc和T-AOC含量升高。发现根系受到盐胁迫较地上部分敏感,为苜蓿耐盐性育种提供了品种选择的依据。
China is the most serious demand country for protein feed, a serious shortage of protein feed will be the key restricting factor in the development of animal husbandry in China.
Alfalfa (Medicago sativa L.) is recognized as the world's largest bank of plant protein and excellent forage due to its high stress resistance, high yield, good quality and good palatability etc., "alfalfa-based dairy industry" is the only way for safe raw milk. As the earliest areas introduction alfalfa of Shaanxi Province, there exist a lot of problems such as varieties confusion, unclear genetic backgrounds and other issues during the process of long-term cultivation, which affecting the industrial development of alfalfa. Based on above reason, the relevant regions of the study 22 alfalfa varieties from domestic and abroad were selected for this study, and RAPD is used to clarify their genetic relationship. At the same time, the agronomic traits, yield traits, and biological characteristics were studied under field conditions for five years, 7 strong heterosis varieties were screened and the flowering habits were observed, which a base work for hybrid and seed production .Finally, the regulatory mechanism of the antioxidant system of alfalfa were studied under salt stress. The results were concluded as follows:
1. The genetic diversity and genetic relationship analysis of 22 alfalfa varieties
22 alfalfa varieties can be clustered into three categories by RAPD:Algonquin and Sandili belong to the first group, Victoria、Total、Queen、Gold Empress、Affinity belong to the same group, the other 15 varieties include 3 domestic varieties were grouped together. Among the 15 varieties, Zhongmu No.1 and Guyuanzihua had relatively close genetic relationship and belonged to the same sub-class,while relatively distant relationship from Guanzhong alfalfa; Haygrazer from Canada had relatively distant relationship from the others and was single grouped, which means that significant heterosis are likely to produced by hybriding it with the other 21 alfalfa varieties.
2. Comparison on the production and quality characteristics of 22 varieties of alfalfa
(1) It was different of the growth stages between varieties, the most obvious difference existed at returning green stage, the earliest and latest gap is 15 days, but with the extension of the growth, the difference was shortened, almost no obvious differences between varieties at flowering period;
(2) The yield of the first cut was higher than those of the 2nd and 3rd cuts in the seeding year. The varieties of Victoria, Total, Sandili, Derfy, Alfalfa 54 and Haygrazer had the better growth performance than those of Guyuanzihua, WL252HQ and Guanzhong alfalfa according to the plant height analysis.
(3) The growth rate of the continuous three harvests showed significant difference (p<0.05) of the 22 alfalfa varieties, varieties of Victoria, Total, Sandili, Derfy, Derby, Alfalfa 54 and Haygrazer showed more advantages. Victoria and Sandili had the highest growth intensity, and the lowest Guanzhong alfalfa showed 6 times lower difference; The varieties that had the fastest regrowth rate include Victoria, Haygrazer, WL324, Alfalfa 54 and WH323ML, of which Victoria and Haygrazer reached 0.8cm / d or more, the worst were still local varieties of Guyuanzihua and Guanzhong alfalfa, only 0.49 cm / d or so;
(4) Variance analysis showed that the fresh and hay yield difference of the 22 alfalfa varieties reached a significant level (p <0.01). The fresh yield of Victoria was significantly higher than the other varieties (p <0.05) except Total, Queen, AmeriGraze, Algonquin, Zhongmu No.1, WL324, Alfalfa 54 and Haygrazer.
(5) WL324 had the highest protein yield of 3216.0 kg/hm2, higher than the lowest Guanzhong alfalfa by 115.0%, and had significantly differences in addition to varieties of Victoria, Alfalfa 54 and Haygrazer.
(6) The Stand height/Plant height(S/P) which describe the erectness of grass showed significantly different among the 22 varieties (p <0.01). The S/P value of good erect Zhongmu No.1, Victoria, WL323HQ, WL232HQ and Derfy were all above 0.75, while the lowest Algonquin, H323ML and Derby only 0.55-0.59.
In summary, 5 advantages varieties of Victoria, Total, Sandili, Affinity, and Haygrazer and two local varieties of Guanzhong alfalfa and Guyuanzihua were screened out for the following hybrid and seed production research.
3. The seed production performance evaluation of 7 alfalfa varieties
(1) The alfalfa flowers generally open at 7:00 to 19:00 in Guanzhong region, the keel petals open peak appeared at 9:00 to 15:00, which was also the peak of insect activity and the best time for alfalfa hybrid;
(2) The correlation analysis showed that the correlation of the florets number and the pod number per inflorescence and seed number per pod were various between different varieties;
(3) The two local varieties showed poor performance of the florets number and the pod number per inflorescence and seed number per pod than the other 5 introduced varieties from abroad.
4. Regulation mechanism of antioxidant system in alfalfa under salt stress
In order to study physiological and biochemical change, seedling of Medicago sativa L.cv. Golden Empress were treated with different NaCl concentration of 0、40、80、120 and 160 mmol/L. Superoxide, MDA, POD, SOD, CAT, Vc and T-AOC of alfalfa leaves and roots were measured. The results showed that the levels of superoxide and MDA increased. Moreover, the activities of POD, SOD and CAT, and antioxidant content of Vc and T-AOC increased in leaves and roots of alfalfa under different NaCl concentration of 0、40、80、120 and 160 mmol/L. Oxidative damage in alfalfa roots may be heavier than in leaves of alfalfa, which provide a theory support for alfalfa salt tolerance breeding.
本研究取得的主要结论有:
1. 22个苜蓿品种的遗传多样性及亲缘关系分析
RAPD标记聚类关系结果表明,22个紫花苜蓿品种可以聚为三大类:引自美国的阿尔岗金和引自荷兰的三得利聚为第一大类,维多利亚、全能、皇后、金皇后、艾菲尼特这5个引自美国的品种聚为第二大类,我国的关中苜蓿、固原紫花、中苜一号和其余品种聚为第三大类。其中,中苜一号和固原紫花苜蓿属于同一亚类,二者之间亲缘关系相对较近,与关中苜蓿之间关系相对较远;加拿大品种放牧者独列一类,与其它紫花苜蓿品种亲缘关系均相对较远,说明用它与其它21个苜蓿品种杂交,均有可能产生明显的杂种优势。
2. 22个苜蓿品种的生产及品质特性比较
(1)不同苜蓿品种的生育时期具有一定的差异,返青期差异最明显,最早和最晚相差达15天,但随着生长的延长其差异间隔天数在缩短,到了开花期,品种间差异几乎不明显;
(2)播种当年头茬苜蓿株高、产量均较低,2茬、3茬较高。株高分析初步可以判断,品种维多利亚、全能、三得利、德福、苜蓿54、放牧者具有较好的生长潜势,而品种关中苜蓿、WL252HQ、固原紫花则相对表现较差;
(3)22个苜蓿品种连续三茬的生长速度差异显著(P<0.05),品种维多利亚、全能、三得利、德福、苜蓿54、放牧者较有优势。品种三得利和品种维多利亚的生长强度最高,关中苜蓿生长强度最低,与品种三得利和品种维多利亚相差近一倍;品种维多利亚、放牧者、WL324、苜蓿54、WH323ML头茬再生速度最快,其中品种维多利亚和品种放牧者达0.8cm/d以上,表现最差的仍为固原紫花和关中苜蓿,其头茬再生速度只有0.49 cm/d左右;
(4)方差分析结果表明,22个苜蓿品种间的鲜干草产量差异均达到极显著水平(P<0.01)。品种维多利亚的鲜草产量除了与全能、皇后、牧歌、阿尔冈金、中苜一号、WL324、苜蓿54、放牧者之间差异不显著外,均显著高出其他品种(P<0.05);
(5)品种WL324(12)单位面积的蛋白质产出量最高,达到3216.0 kg/hm2,比最低的关中苜蓿高出115.0%,除了与品种维多利亚、苜蓿54、放牧者差异不显著外,与其他品种差异均显著;
(6)品种间草株比具有极显著差异(P<0.01)。中苜一号、维多利亚、WL323HQ、WL232HQ、德福草株比最高,都在0.75以上,直立性最好;品种阿尔冈金、WH323ML、德宝最低,只有0.55-0.59。
综上,维多利亚、全能、三得利、爱非利特和放牧者5个优势品种和两个地方品种关中苜蓿、固原紫花被筛选出来并做为下一步杂交和种子生产习性研究的材料。
3. 7个紫花苜蓿品种的种子生产性能评价
(1)关中地区紫花苜蓿一般在7:00~19:00小花开放,9:00~15:00为龙骨瓣开放高峰期,是昆虫活动的高峰期,也是进行苜蓿杂交的最佳时期;
(2) 7个紫花苜蓿品种单个花序小花数、单个花序结荚数和单个荚内种子数相关性分析结果显示各指标之间的相关性因品种而异;
(3)关中苜蓿和固原紫花2个紫花苜蓿品种的单个花序小花数、单个花序结荚数和单个荚内种子数均不及国外引进的三得利、爱非尼特、放牧者、全能和维多利亚5个紫花苜蓿品种。
4.盐胁迫下紫花苜蓿抗氧化系统的调节机制
为研究盐胁迫下紫花苜蓿对逆境生理生化的调节机制,以紫花苜蓿幼苗为研究材料,采用不同NaCl浓度(0、40、80、120和160 mmol/L)进行盐胁迫处理,对其幼苗叶片和根部的超氧阴离子自由基( O_2~(·-) ),膜脂过氧化产物丙二醛(MDA),以及抗氧化系统酶中的过氧化物酶(POD),超氧化物歧化酶(SOD),过氧化氢酶(CAT)和非酶抗氧化剂Vc和总抗氧化能力(T-AOC)等指标进行了测定。研究结果表明,随着NaCl盐浓度增加,超氧阴离子自由基和MDA积累增加,抗氧化物酶POD、SOD和CAT活性也不断增加,同时非酶抗氧化剂Vc和T-AOC含量升高。发现根系受到盐胁迫较地上部分敏感,为苜蓿耐盐性育种提供了品种选择的依据。
China is the most serious demand country for protein feed, a serious shortage of protein feed will be the key restricting factor in the development of animal husbandry in China.
Alfalfa (Medicago sativa L.) is recognized as the world's largest bank of plant protein and excellent forage due to its high stress resistance, high yield, good quality and good palatability etc., "alfalfa-based dairy industry" is the only way for safe raw milk. As the earliest areas introduction alfalfa of Shaanxi Province, there exist a lot of problems such as varieties confusion, unclear genetic backgrounds and other issues during the process of long-term cultivation, which affecting the industrial development of alfalfa. Based on above reason, the relevant regions of the study 22 alfalfa varieties from domestic and abroad were selected for this study, and RAPD is used to clarify their genetic relationship. At the same time, the agronomic traits, yield traits, and biological characteristics were studied under field conditions for five years, 7 strong heterosis varieties were screened and the flowering habits were observed, which a base work for hybrid and seed production .Finally, the regulatory mechanism of the antioxidant system of alfalfa were studied under salt stress. The results were concluded as follows:
1. The genetic diversity and genetic relationship analysis of 22 alfalfa varieties
22 alfalfa varieties can be clustered into three categories by RAPD:Algonquin and Sandili belong to the first group, Victoria、Total、Queen、Gold Empress、Affinity belong to the same group, the other 15 varieties include 3 domestic varieties were grouped together. Among the 15 varieties, Zhongmu No.1 and Guyuanzihua had relatively close genetic relationship and belonged to the same sub-class,while relatively distant relationship from Guanzhong alfalfa; Haygrazer from Canada had relatively distant relationship from the others and was single grouped, which means that significant heterosis are likely to produced by hybriding it with the other 21 alfalfa varieties.
2. Comparison on the production and quality characteristics of 22 varieties of alfalfa
(1) It was different of the growth stages between varieties, the most obvious difference existed at returning green stage, the earliest and latest gap is 15 days, but with the extension of the growth, the difference was shortened, almost no obvious differences between varieties at flowering period;
(2) The yield of the first cut was higher than those of the 2nd and 3rd cuts in the seeding year. The varieties of Victoria, Total, Sandili, Derfy, Alfalfa 54 and Haygrazer had the better growth performance than those of Guyuanzihua, WL252HQ and Guanzhong alfalfa according to the plant height analysis.
(3) The growth rate of the continuous three harvests showed significant difference (p<0.05) of the 22 alfalfa varieties, varieties of Victoria, Total, Sandili, Derfy, Derby, Alfalfa 54 and Haygrazer showed more advantages. Victoria and Sandili had the highest growth intensity, and the lowest Guanzhong alfalfa showed 6 times lower difference; The varieties that had the fastest regrowth rate include Victoria, Haygrazer, WL324, Alfalfa 54 and WH323ML, of which Victoria and Haygrazer reached 0.8cm / d or more, the worst were still local varieties of Guyuanzihua and Guanzhong alfalfa, only 0.49 cm / d or so;
(4) Variance analysis showed that the fresh and hay yield difference of the 22 alfalfa varieties reached a significant level (p <0.01). The fresh yield of Victoria was significantly higher than the other varieties (p <0.05) except Total, Queen, AmeriGraze, Algonquin, Zhongmu No.1, WL324, Alfalfa 54 and Haygrazer.
(5) WL324 had the highest protein yield of 3216.0 kg/hm2, higher than the lowest Guanzhong alfalfa by 115.0%, and had significantly differences in addition to varieties of Victoria, Alfalfa 54 and Haygrazer.
(6) The Stand height/Plant height(S/P) which describe the erectness of grass showed significantly different among the 22 varieties (p <0.01). The S/P value of good erect Zhongmu No.1, Victoria, WL323HQ, WL232HQ and Derfy were all above 0.75, while the lowest Algonquin, H323ML and Derby only 0.55-0.59.
In summary, 5 advantages varieties of Victoria, Total, Sandili, Affinity, and Haygrazer and two local varieties of Guanzhong alfalfa and Guyuanzihua were screened out for the following hybrid and seed production research.
3. The seed production performance evaluation of 7 alfalfa varieties
(1) The alfalfa flowers generally open at 7:00 to 19:00 in Guanzhong region, the keel petals open peak appeared at 9:00 to 15:00, which was also the peak of insect activity and the best time for alfalfa hybrid;
(2) The correlation analysis showed that the correlation of the florets number and the pod number per inflorescence and seed number per pod were various between different varieties;
(3) The two local varieties showed poor performance of the florets number and the pod number per inflorescence and seed number per pod than the other 5 introduced varieties from abroad.
4. Regulation mechanism of antioxidant system in alfalfa under salt stress
In order to study physiological and biochemical change, seedling of Medicago sativa L.cv. Golden Empress were treated with different NaCl concentration of 0、40、80、120 and 160 mmol/L. Superoxide, MDA, POD, SOD, CAT, Vc and T-AOC of alfalfa leaves and roots were measured. The results showed that the levels of superoxide and MDA increased. Moreover, the activities of POD, SOD and CAT, and antioxidant content of Vc and T-AOC increased in leaves and roots of alfalfa under different NaCl concentration of 0、40、80、120 and 160 mmol/L. Oxidative damage in alfalfa roots may be heavier than in leaves of alfalfa, which provide a theory support for alfalfa salt tolerance breeding.
引文
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