两种冷季型草坪草的EMS诱变及突变体筛选

Ethyl methanesulfonate mutagenesis and mutant screening of two cold-season turfgrasses

为确定甲基磺酸乙酯(EMS)诱变多年生黑麦草‘爱神特’(Lolium perenne‘Accent’)和高羊茅‘赤道’(Festuca arundinacea‘Equator’)种子的最佳条件,获得性状稳定的突变材料和后代。利用不同浓度EMS在不同时间下诱变两个草坪草种,根据半致死浓度确定诱变组合。通过形态学、SCoT分子标记和生理指标测定筛选M1和M2代突变苗。结果表明:使用0.8%EMS溶液处理两个草坪草种36 h后播种,发芽率接近50%,诱变效果最佳。以该参数大规模诱变后,M1代两个草坪草种诱变株移栽成活933株,再经短期干旱胁迫,共筛选出18株耐旱单株。对M2代诱变株形态学分析发现,有叶面狭窄型和根长增长型两种突变类型。利用SCoT分子标记技术验证M2代诱变株发现,筛选的5条SCoT标记引物对20份草坪草样品扩增后,多态性比率为84.09%。其中SCoT-35引物扩增产物条带最为清晰稳定,诱变株与野生型差异明显。M2代三叶期变异单株经干旱、高温胁迫处理后,丙二醛含量、脯氨酸含量及相对电导率呈升高趋势,叶绿素含量呈下降趋势。最终得到两株多年生黑麦草耐旱突变体A5和A8,1株高羊茅耐旱突变体B9;两株多年生黑麦草耐热突变体A8和B9,1株高羊茅耐旱耐热突变体B9。本研究结果为草坪草抗性品种选育提供了帮助。

This study aimed to determine the optimal mutagenesis conditions for ethyl methane sulfonate(EMS)mutagenesis treatment of Lolium perenne‘Accent’and Festuca arundinacea‘Equator’seeds and to obtain mutant material and progeny with stable traits.Seeds of both turfgrasses were mutagenized using different EMS concentrations and treatment times,and mutagenesis combinations were determined according to the semi-lethal concentration.The best mutagenic effect was achieved by treating two turfgrass seeds with 0.8%EMS solution for 36 h and then sowing them with germination rates close to 50%.After large-scale mutagenesis with these parameters,933 mutant plants of two M1 generation turfgrass species were transplanted and survived,and a series of 18 drought-tolerant monocultures were screened after short-term drought stress.Morphological analysis of the M2 generation mutant plants revealed two mutation types:foliar narrowing and root length growth.Amplification of 20 turfgrass samples with five SCoT marker primers showed a polymorphism ratio of 84.09%.Among them,the SCoT-35 primer amplification product had the clearest and most stable bands,and the difference between the mutant and wild type was obvious.After the M2 generation of three-leaf stage mutant plants was treated with drought and high temperature stress,the malondialdehyde content,proline content,and relative conductivity all tended to increase,and the chlorophyll content tended to decrease.Two L.perenne drought-tolerant mutants A5 and A8;one F.arundinacea drought-tolerant mutant B9;two heat-tolerant mutants A8 and B9;and one drought and heat-tolerant mutant B-9 were finally obtained.The results of this study provide help for the breeding of resistant turfgrass varieties.