【作者】 马艳红；

【导师】 于卓；

【作者基本信息】 内蒙古农业大学 ， 作物栽培学与耕作学， 2007， 博士

【摘要】 小麦族多年生禾草的多数种具有耐瘠薄、抗逆性强、优质高产等特性,是禾草及麦类作物品种改良的宝贵基因库。为创造结合双亲优良特性的饲草新种质,我们将引自北美洲草原具高产优质特性的加拿大披碱草分别与产于内蒙古草原抗逆性强的野大麦、披碱草、圆柱披碱草进行种属间远缘杂交,获得了高度不育的杂种F1代。本研究利用秋水仙碱诱导染色体加倍法和回交法克服这3个杂种的不育性,并从形态及生物学、细胞学、同工酶及AFLP不同水平分别对加拿大披碱草—野大麦染色体加倍后代、加拿大披碱草—披碱草F1和加拿大披碱草—圆柱披碱草F1及其BC1代进行了鉴定分析。主要结果如下:1.秋水仙碱处理三倍体加拿大披碱草—野大麦F1分蘖苗的适宜浓度和时间范围分别为0.15%～0.20%和12～24h,获得染色体加倍植株17个,并在田间发现了16个染色体自然加倍株丛。2.加拿大披碱草—野大麦染色体自然加倍植株及其加倍F1代、秋水仙碱加倍植株的PMCMⅠ染色体构型分别为0.04Ⅰ+20.96Ⅱ、0.03Ⅰ+20.98Ⅱ和0.03Ⅰ+20.97Ⅱ,花粉可育率分别为84.97%、94.08%和90.21%,自然结实率分别为77.51%、83.87%和80.13%,育性较高。3.加拿大披碱草—野大麦染色体加倍植株及其后代的生长势强,生育期均为147d,开花持续期和果后营养期长,枯黄期晚,株高、叶长、叶宽、穗长、穗宽等形态特征明显优于杂种F1,继承了父本野大麦分蘖性和耐盐性强的特性。4.加拿大披碱草—野大麦染色体加倍植株及其加倍F1在同一发育阶段的EST、POD和SOD 3种同工酶酶谱表征具有一致性,而在不同发育阶段同工酶酶谱表征存在明显差异,可作为染色体加倍后代育性恢复鉴定的遗传标记。5.用19个AFLP适宜引物对加拿大披碱草—野大麦染色体加倍植株及其加倍F1～F4、亲本及杂种F1扩增出的多态性位点比率为84.60%;聚类分析表明,亲本加拿大披碱草与野大麦遗传差异最大,杂种F1与♀加拿大披碱草遗传差异相对较小,染色体加倍植株及其加倍F1～F4代间的遗传差异最小。6.加拿大披碱草—披碱草F1生长速度明显超过双亲、加拿大披碱草—圆柱披碱草F1生长速度介于双亲之间,其生育期分别为139d和132d,株高分别为150.05cm和143.67cm,株型均偏向♀加拿大披碱草,叶片长而宽大,叶量丰富,穗子大,分蘖能力强。7.加拿大披碱草—披碱草F1、加拿大披碱草—圆柱披碱草F1均为五倍体,其PMCMⅠ染色体构型分别为19.04Ⅰ+7.45Ⅱ+ 0.28Ⅲ+0.02Ⅳ和5.82Ⅰ+14.31Ⅱ+ 0.15Ⅲ,染色体配对行为不规则是造成杂种不育的重要原因。8.用13个AFLP适宜引物对加拿大披碱草—披碱草F1、加拿大披碱草—圆柱披碱草F1及其双亲扩增的多态性位点比率为83.56%,各材料间的AFLP指纹图谱差异明显,可作为杂种F1与其各自亲本识别的分子依据;聚类分析显示,亲本加拿大披碱草与披碱草的遗传差异比与圆柱披碱草的差异大,2个杂种F1与各自父本的遗传差异相对较小。9.建立了加拿大披碱草—披碱草F1、加拿大披碱草—圆柱披碱草F1组培再生体系,即以MS为基本培养基,愈伤组织诱导、继代、幼苗分化、生根的最适培养基浓度组合分别为:400mg/L CH +3.0mg/L 2,4-D、400mg/L CH + 1.0mg/L 6BA+0.5 mg/L 2,4-D、400mg/L CH +3.0mg/L 6BA +0.5mg/L NAA和0.5mg/L NAA + 0.5mg /L IBA;在此基础上,用秋水仙碱诱导2个杂种F1的愈伤组织各获得4个变异植株。10.获得[加拿大披碱草×披碱草]×加拿大披碱草BC1植株4个、[加拿大披碱草×圆柱披碱草]×加拿大披碱草BC1植株3个,其染色体数目均趋向加拿大披碱草。更多还原

【Abstract】 The majority species of perennial Triticeae grasses had the characters of barren tolerance, stronger tress tolerance, excellent quality and high yield, which were precious gene bank in crop varietal improvement. In order to create new forage grass germplasms of parental superior characters, wide crosses between Elymus canadensis of excellent quality and high yield from north America (2n=4x=28, genome SSHcHc) and Hordeum brevisubulatum(2n=4x=28,genome H1H1H2H2), E. dahuricus(2n=6x=42, genome SpSpHpHpYpYp), E. cylindricus(2n=6x=42, genome SoSoHoHoYoYo) of strong drought resistance and tillering from China were made respectively, and 3 intergeneric and interspecific hybrids F1 which were completely sterile were successfully gotten. The sterility of hybrids F1 were restored with the methods of inducing chromosome doubling with colchicines and by backcross, and Elymus canadensis- Hordeum brevisubulatum doubling plants and its descendants, E. canadensis- E. dahuricus hybrid F1 and backcross 1(BC1), E. canadensis- E. cylindricus hybrid F1 and BC1 were identified and analysed based on morph-biology, cytology, isozyme and AFLP maker. The main results as follows:1. The suitable colchicine concentration and treatment time for the tillering plants of E. canadensis- H. brevisubulatum triploid hybrid F1 were 0.15%～0.20% and 12～24h respectively, and 17 chromosome doubling plants were got by colchicine induction, and 16 natural chromosome doubling clumped plants were found in field.2. The average chromosome pairing configuration at metaphaseⅠin pollen mother cells (PMCMⅠ) of the natural chromosome doubling plant, natural chromosome doubling F1 and cochicines induced chromosome doubling plant of triploid hybrid F1 were 0.04Ⅰ+ 20.96Ⅱ, 0.03Ⅰ+ 20.98Ⅱa nd 0.03Ⅰ+ 20.97Ⅱrespectively, their pollen fertility rate were 84.97%, 94.08% and 90.21% respectively, the seed set were 77.51%, 83.87% and 80.13% respectively, and had higher fertility.3. The growth vigour of chromosome doubling plant and its descendants were stronger, their growth periods were 147d, and had the characters of long flowering duration, long growth period after fructification and late withering, and their morpholo- gical characters of plant height, leaf length and width, ear length and width et al. exceled hybrid F1, and had the strong tillering vigor and capability of salt tolerance from paternal H. brevisubulatum.4. The E. canadensis-H. brevisubulatum chromosome doubling plant and doubling F1 had uniformity and genetic stability in the isozyme zymograms token(bands number, locus and intensity) of esterase (EST), proxidase (POD) and superoxide dismutase (SOD) isozyme at the same growth-development stage, but they were significantly different from the hybrid F1 and their parents in isozyme band patterns, the isozyme zymograms can be used as genetic marker to identify fertility recovering of chromosome doubling plant of hybrid.5. The percentage of polymorphism loci of AFLP markers with 19 primer combina- tions to E. canadensis- H. brevisubulatum chromosome doubling plant and doubling F1, their parents and hybrid F1 was 84.60%; UPGMA cluster analysis based on AFLP markers indicated that the genetic diversity between E. canadensis and H. brevisubula- tum was maximal, and the genetic diversity between hybrid F1 and♀E. canadensis was lesser, and the genetic diversity among chromosome doubling plant and doubling F1～F4 was the least.6. The growth rate of E. canadensis- E. dahuricus hybrid F1 was more than its parents, and the growth rate of E. canadensis- E. cylindricus hybrid F1 was in the middle of parents, their growth periods were 139d and 132d respectively, plant height were 150.05cm and 143.67cm, plant type was similar to E. canadensis, the size and number of leaves were bigger, the ear of F1 hybrids was bigger than their parents, had the strong tillering vigor.7. E. canadensis- E. dahuricus hybrid F1 and E. canadensis- E. cylindricus hybrid F1 were pentaploid (2n=5x=35), the average chromosome pairing configurations at PMCMⅠwere 19.04Ⅰ+7.45Ⅱ+0.28Ⅲ+0.02Ⅳand 5.82Ⅰ+14.31Ⅱ+ 0.15Ⅲrespec- tively, the important reasons why hybrid F1 was sterile was that chromosome pairing behavior was irregular.8. The percentage of polymorphism loci of AFLP markers with 13 primer combina- tions to E. canadensis- E. dahuricus hybrid F1, E. canadensis- E. cylindricus hybrid F1 and their parents with was 83.56%, and they were significantly different from the hybrid F1 and their parents in AFLP fingerprint, the fingerprint can be used as molecular marker to identify hybrid F1 and their parents; the cluster analysis indicated that the genetic diversity between E. canadensis and E. dahuricus was bigger than that of E. canadensis and E. cylindricus, and the genetic diversity between two hybrids F1 and their paternal was lesser.9. The system of tissue culture and plant regeneration two hybrids F1 of E. canadensis- E. dahuricus and E. canadensis- E. cylindricus were established. MS medium as the basic medium, the concentration combination of optimal medium on callus induction, callus subculture, plantlets differentiation and rooting were 400mg/L CH +3.0mg/L 2,4-D、400mg/L CH + 1.0mg/L 6BA+0.5 mg/L 2,4-D、400mg/L CH +3.0mg/L 6BA +0.5mg/L NAA and 0.5mg/L NAA + 0.5mg /L IBA respectively. On the system of tissue culture, eight variation plants were achieved by colchicine induction callus of two hybrids F1.10. Four BC1 lines of [E. canadensis×E. dahuricus]×E. canadensis and three BC1 lines of [E. canadensis×E. cylindricus ]×E. canadensis were obtained, their choromo- some number trended to E. canadensis.更多还原