不同培肥措施下种植制度及撂荒对土壤微生物量碳氮的影响

以黄土高原南部半湿润易旱区已进行17年的田间定位试验为研究对象,研究了不同培肥措施(不施肥、施用氮磷钾及氮磷钾与有机肥配合施用)下两种种植制度(一年1熟及一年两熟)和撂荒对土壤微生物量碳、氮(SMBC、SMBN)及可溶性有机碳、氮(SOC、SON)等含量的影响。结果表明,与一年1熟的小麦-休闲种植制度相比,一年两熟小麦-玉米轮作提高了0~10cm土层SMBC、SMBN、有机碳(TOC)、全氮(TN)和土壤SOC、SON的含量,而对10~20cm土层上述测定指标影响不大。与不施肥(CK)或单施化肥处理(NPK)下小麦-休闲和小麦-玉米轮作方式相比,撂荒处理显著提高了0~10cm土层各测定指标的含量。不同培肥措施相比,氮磷钾配施有机肥显著提高了0~10cm、10~20cm土层SMBC、SMBN含量;NPK处理0~10cm土层SMBN含量显著增加,10~20cm土层SMBN和0~10cm、10~20cm土层SMBC含量增加但未达显著水平。不同培肥措施和种植制度对SMBC/TOC和SMBN/TN的比例无明显影响。

宁夏南部山区秸秆覆盖对春玉米水分利用及产量的影响

为研究秸秆覆盖在宁南山区对春玉米生长及其土壤水分的影响,在宁夏彭阳县对三种不同秸秆覆盖处理方式进行了试验研究。对比分析了不同覆盖措施下春玉米的生长状况、产量及水分利用效率变化的关系。结果表明:采用秸秆覆盖方式可使春玉米的株高、穗位高、穗长、生物产量及经济产量等指标得到显著提高,土壤的蓄水保墒性能增强,水分利用效率提高,增产增效明显;与传统方式相比,整秸秆覆盖可使春玉米的产量及水分利用效率分别提高3.5%及16.5%。

沃特和PAM施用方式对土壤水分及玉米生长的影响

为了鉴别沃特保水剂和PAM不同施用方式的施用效果,在陕北黄土丘陵沟壑区开展了撒施、沟施、穴施对土壤水含量和玉米生长影响的田间试验。结果表明不同施用方式提高了0～10cm土层土壤水含量,而30～40cm土层土壤水含量则随降水量的多少呈现出不同的规律性。随着玉米的生长,不同施用方式影响的土层深度逐渐加深,从三叶期0～50cm土层增加到成熟期的0～200cm土层,土壤水含量表现为沟施、穴施高于撒施,撒施高于对照。3种施用方式对玉米出苗无显著影响,但均提高了玉米生物量、籽粒产量、水分利用效率,降低了玉米的耗水量,其中沟施、穴施的效果强于撒施。沃特、PAM相同施用方式对土壤水含量和玉米生长无显著影响,且沟施和穴施之间无显著性差异,但沃特单位施用量单位面积的增产量高于PAM。玉米生产中,沃特、PAM应以沟施或穴施为主。

长期施氮磷化肥对不同种植体系土壤有效硫累积研究

以22 a定位试验为基础,在黄土高原旱地研究了长期施用氮磷化肥,对不同种植体系土壤有效硫在剖面上分布与累积状况。结果表明,在60-80 cm土层各处理出现第一个累积峰,累积峰值为:玉米-小麦(2 a)+糜子轮作27.07 mg/kg,豌豆-小麦(2 a)+糜子轮作25.42 mg/kg,小麦(2 a)+糜子-玉米轮作24.23 mg/kg,豌豆-小麦(2 a)+玉米轮作22.61 mg/kg,小麦连作16.56 mg/kg,红豆草-小麦(2 a)轮作15.14 mg/kg;在120-180 cm土层又出现有效硫的第二个累积峰,累积峰值为:小麦(2 a)+糜子-玉米轮作34.20 mg/kg,豌豆-小麦(2 a)+糜子轮作32.16 mg/kg,豌豆-小麦(2 a)+玉米轮作31.00 mg/kg,红豆草-小麦(2 a)轮作30.32mg/kg,玉米-小麦(2 a)+糜子轮作29.16 mg/kg,小麦连作26.22 mg/kg。0-200 cm土层有效硫总累积量玉米-小麦+糜子轮作高达559.64 kg/hm2,其次是小麦+糜子-玉米轮作为538.88 kg/hm2,豌豆-小麦+糜子轮作为514.34 kg/...

新型土壤改良剂对夏玉米生理生长特性的影响

通过大田玉米试验,验证新型土壤改良剂对夏玉米生育期土壤水分、紧实度及玉米生理生长特性的影响。结果表明,施用改良剂PJG和PFL夏玉米全生育期平均株高、叶面积分别高于对照20.7%、19.75%和51.88%、72.37%;2种改良剂对干物质积累的影响存在差异,影响效果依次为茎干重>叶干重>根干重。夏玉米光合速率和叶绿素含量受土壤改良剂影响较大,PJG和PFL分别高于对照29.96%、24.48%和73.36%、68.53%。在0~10 cm土层内,施用PJG和PFL后土壤紧实度分别低于对照44.44%和42.91%。施用改良剂PJG后,0~20 cm土层土壤含水量维持在田间持水量的77.9%左右,未施用改良剂土壤,夏玉米生育期表层土壤含水量起伏变化较大。土壤改良剂PJG在夏玉米的施用效果略好于PFL。

秸秆覆盖量对夏玉米产量影响的试验研究

通过对夏玉米在露天和干旱棚控水2种土壤水分状况下生长状况的观测,发现秸秆覆盖夏玉米有明显增产效应,作物水分利用效率均得到了不同程度提高。通过对0、3000、6000、9000、12000 kg/hm2五种处理下玉米状况的对比,得出6000 kg/hm2为渭河平原地区最适宜夏玉米田秸秆覆盖量。在6000 kg/hm2覆盖量下,露天小区夏玉米可增产5.61%。试验证明,渭河平原地区秸秆覆盖夏玉米,增产潜力大,是一种值得推广的玉米增产措施。

半干旱区施氮和灌溉条件下覆膜对春玉米产量及氮素平衡的影响

以典型半干旱区干湿砂质新成土(Ust-Sandic Entisols)为供试土壤进行田间试验,研究地膜覆盖、施氮及补充灌水量对春玉米(Zea maysL.)产量、土壤矿质氮(NO3--N和NH4+-N)及氮素平衡的影响。结果表明,0—100 cm土体范围内,随着土层加深,播前和收获后土壤NO3--N含量呈降低趋势,NH4+-N有所增加,但变幅不大;总矿质氮量(NO3--N和NH4+-N)表现为下降。说明地膜覆盖和施氮并没有使NO3--N深层累积量增加,这可能与土壤本身供氮能力严重不足有关。与不施氮相比,施氮各处理氮肥表观损失量增加;与不覆膜相比,作物氮素累积量比不覆膜显著增加(P<0.05)。在低灌(80 mm)覆膜和高灌(160 mm)覆膜条件下,玉米的氮肥利用率均比不覆膜提高了18.8%,说明覆膜低灌在相同施氮条件下,可节约80 mm灌水。但低灌(80 mm)与高灌(160 mm)不覆膜间氮肥利用率差异不显著,表明在相同施氮条件下,覆膜可有效提高氮肥利用率,减少氮素损失。综合考虑子粒产量和氮肥利用率,"覆膜+补灌80 mm+施氮90 kg/hm2"可能为本试验条件下较优的栽培模式。

夏玉米冠层叶片表观自由空间的差异研究

叶片自由空间在环境与冠层养分交换间具有重要作用,是目前植物营养学研究的重要领域之一。通过盆栽试验,以不同株型夏玉米为试材,研究了不同施氮水平(N 0、0.15和0.30 g/kg)下玉米冠层叶片表观自由空间(AFS)的差异。结果表明,不同生育期叶片AFS差异极显著(P<0.01),表现为随生育期推进,叶片AFS、生物量和全氮含量均逐渐降低;除成熟期,其它各生育期品种与施氮水平对叶片AFS的影响均存在显著的交互作用。施氮对叶片生物量的影响不显著,但施氮后叶片全氮含量显著增加。不同施氮水平下,植株冠层叶片AFS也存在显著差异,中量施氮处理(即N 0.15 g/kg)植株叶片AFS值(9.49)明显高于不施氮处理(9.03),但随施氮量进一步增加,叶片AFS下降,施氮量为N 0.30 g/kg时,叶片AFS为8.62;不同施氮水平各生育期不同叶层间叶片AFS、生物量和全氮含量差异显著(P<0.05)。不同品种间叶片全氮含量和AFS存在显著差异(P<0.05),以紧凑型品种陕单902叶片AFS最大,平均为9.24,显著高于其它品种;中间型品种农大108与平展型品种陕单9号间差异不显著,二者平均分别为9.06和8.85...

玉米根系活力杂种优势及其与光合特性的关系

对大田不同水分处理条件下玉米杂交种及其亲本在经济产量形成期的根系活力及其衰退的杂种优势进行了研究,并分析了根系活力与叶片光合特性之间的关系。结果表明,水分亏缺显著降低了玉米的根系活力;充分供水和水分亏缺条件下,玉米杂交种根系活力都具有显著的杂种优势;杂交玉米种根系活力衰退速率具有显著的负优势,抽雄后灌水能够显著降低玉米根系活力衰退速率。杂交玉米种在抽雄期各水分处理下的光合速率显著大于亲本;玉米根系活力与光合速率之间具有显著的正相关关系,随着根系活力的增高,玉米光合速率、气孔导度和蒸腾速率均增高。因此,杂交种较高的抗旱性是根冠共同作用的结果。在玉米育种和栽培中,要充分地考虑到根系对地上部生理活性的影响。

聚丙烯酸钠对3种土壤持水能力及作物产量的影响

在离心机模拟不同水吸力条件下,研究了聚丙烯酸钠(sodium polyacrylate记作SP)5种使用浓度(占干土质量0,0.01%,0.08%,0.2%与1%)对3种土壤(砂土、壤土、黏土)持水能力的影响;采用大田试验研究了地表撒施2 g/m2SP对冬小麦与下季玉米产量及WUE影响。结果表明:3种土壤在0.01 MPa至1.5 MPa水吸力下的持水能力随着SP用量的增加而增加,砂土的作用效果较壤土、黏土更显著;3种土壤适宜浓度为0.08%~0.2%,最佳用量为0.2%,此用量条件下砂土、壤土、黏土的最大毛管持水量分别较对照增加了138.61%,7.22%,62.70%;不灌水条件下,SP处理较不施用SP冬小麦增产4%,WUE增加5.7%,灌浆期灌水28.5 mm条件下SP处理较不施用SP增产1%,WUE降低1%;SP处理的玉米产量较对照降低0.5%,WUE提高3%,效果明显低于对冬小麦效果。

下辽河平原玉米农田水肥交互作用研究

利用持续16年的长期田间试验与气象观测资料，对下辽河平原不同降水年景作物对施肥的增产反应及水分对肥料增产的促进作用、不同水肥条件下玉米对养分的吸收与分配以及玉米农田土壤养分收支进行了研究；并估算了该地区水肥交互作用对作物增产的贡献；同时，利用沈阳地区近50年气象与玉米单产历史资料，建立趋势—随机预测模型，以预测该地区不同气候年景玉米产量。 研究结果表明，该地区玉米蒸散耗水与其生育期内降水之间关系密切，且不同的施肥处理对玉米消耗水分量影响不大。过少或过多的降水均会降低玉米产量；在高肥力水平下平水年（降水400～550mm）玉米产量最高，水肥交互作用对作物增产的贡献亦达最大，可占总增产量的44.9%。 玉米体内养分浓度及养分在籽实与秸秆中的分配比例明显受施肥、降水及作物生长状况的影响。在平水年条件下，养分在籽实内累积的比例最高；同时，作物生长状况愈好，形成单位籽实产量所需养分量有减少的趋势，有利于提高作物体内养分的转化利用效率。 施肥条件差异可使玉米收获移出农田的养分量相差2～3倍。水分条件对养分移出量的作用不如施肥明显；N肥施用加剧了土壤P的赤字，而N、P肥施用可加剧土壤K收支赤字；水分条件的改善也会明显加快养分收支赤字过程。 通过对沈阳地区玉米单产与气象因子的分析，建立趋势—随机预测模型，发现热量因素对玉米产量形成有着重要作用；而养分均衡供给可提高玉米对不利气象条件的耐抗能力，具有稳产作用；同时，利用长期定位试验结果对模型加以验证，发现模型预测结果与长期定位试验结果非常吻合。

松辽平原农田土壤碳氮磷分布特征研究

本文对东北地区松辽平原不同纬度农田土壤碳氮磷剖面分布特征进行比较研究，从北到南依次采集了黑土区的海伦、哈尔滨、德惠、公主岭和棕壤区的昌图、沈阳、大石桥玉米地土壤样品。所得主要结论如下： 各样点土壤有机碳含量随土层深度的增加而下降。海伦、哈尔滨和公主岭样点40～60cm土层土壤有机碳含量及其储量显著低于0～40cm土层；海伦、哈尔滨、德惠、公主岭和昌图样点土壤水溶性有机碳表现出随深度增加先升高后降低，在沈阳和大石桥样点土壤水溶性有机碳表现出随深度而下降的趋势；各样点0～20cm土层土壤微生物量碳含量高于20～40cm土层。典型黑土区海伦点0～100cm的SOC储量为213.4t•hm-2, 棕壤区昌图、沈阳、大石桥样点分别为69.9、87.9和73.4t•hm-2，海伦点SOC储量是棕壤区三样点的3倍左右。 土壤全氮、碱解氮、硝态氮及氮储量随剖面深度增加而下降。德惠点在20～40cm土层、沈阳点在40～60cm土层、昌图点在60～80cm土层的全磷含量最低；其他样点土壤全磷、有机磷含量和磷储量总体上呈现随土层深度增加而下降的趋势。黑土区样点土壤有机磷含量在40cm以下各土层迅速下降，而棕壤区各样点20cm以下各土层差异不显著。除公主岭和大石桥点外, 其他各样点土壤Olsen-P含量在0～20cm 土层显著高于20～40cm土层。 土壤有机碳、全氮、碱解氮、全磷和有机磷含量随纬度增加而增加。营养元素在纬度上的分异主要受成土母质、气候条件等自然因素影响，施肥、耕作等人为活动对表层土壤营养元素分布的影响较大。除土壤水溶性有机碳外，土壤碳、氮和磷之间及其与其他基本理化性质间均存在显著的相关关系。

乙草胺对玉米根际微生物和作物抗氧化酶活性的影响

乙草胺作为我国应用最多的三大除草剂之一，被广泛地应用于玉米、花生、大豆等农田中。作为外源有机污染物，乙草胺在施入土壤后，对作物构成一定的逆境胁迫。作物在逆境胁迫下，自身会发生相应的改变抵御迫害，其中根际、活性氧清除系统、除草剂的代谢酶等在作物抵御乙草胺等除草剂的迫害中起着十分重要的作用。 本文以我国重要的粮食作物-玉米作为供试作物，在田间实验条件下研究了低（田间用量）、中（田间用量的2倍）、高（田间用量的5倍）三个浓度乙草胺处理下，乙草胺在玉米根际和非根际环境中的残留特征，并探讨了玉米根际微生物和玉米超氧化物歧化酶、过氧化物酶、还原性谷胱甘肽、谷胱甘肽-S-转移酶对乙草胺施用的响应特征。 研究表明：（1）乙草胺在土壤中的降解迅速，并且在根际的降解得到强化，随着乙草胺的施用浓度增加，其在土壤中的最终残留量增大。（2）由于根际效应的存在，根际微生物量、真菌和细菌数量都显著高于非根际土壤，且根际和非根际土壤、高浓度处理和低、中浓度处理土壤间微生物群落结构明显不同。乙草胺各处理对根际微生物量有先抑制后刺激的作用，对根际细菌尤为显著，抑制作用随乙草胺浓度的增加而加强。乙草胺对本体土壤的微生物量影响较复杂，高浓度对本体土壤微生物量主要表现为抑制作用，而低、中浓度则是抑制和刺激作用交替出现，低浓度乙草胺对非根际土壤微生物的抑制作用未达到显著水平。乙草胺对对非根际真菌的抑制程度大于根际真菌。虽然各处理的微生物量都能恢复到正常水平，但非根际的恢复比根际需要更长的时间。土壤中细菌和真菌对乙草胺响应存在非同步性，并且群落变化具有一定的补偿作用。（3）除草剂乙草胺对玉米过氧化物酶（POD）的活性影响较小，POD不能作为乙草胺污染的生物标记物；超氧化物歧化酶（SOD）对乙草胺的污染有一定的指示作用，但其对乙草胺的敏感性不如谷胱甘肽-S-转移酶（GST）和还原性谷胱甘肽（GSH）高。低、中浓度乙草胺处理使玉米GST活性和GSH含量先升高后降低，高浓度抑制GST活性，降低GSH的含量。GST和GSH对乙草胺的响应非常敏感，可以作为评价除草剂对作物安全性评价的重要指标。 5倍田间用量乙草胺处理虽然对玉米产生了一定的影响，但玉米经一个月左右能恢复正常生长，说明乙草胺对玉米的安全性较高。但由于乙草胺在土壤中的最终残留量与初始浓度密切相关，且对土壤（根际和非根际）微生物群落会产生一定影响，所以应严格控制乙草胺的用量，以降低对土壤环境、水体环境和人类生活健康的风险性。

同源四倍体水稻三系亲本的遗传差异、细胞遗传学比较研究及低直链淀粉含量突变体Wx基因序列分析

同源四倍体水稻（2N=4X=48,AAAA）是由二倍体水稻（2N=2X=24,AA）通过秋水仙素诱导染色体加倍后得到的新品系，具有优良的抗病性以及较高的蛋白质含量。因此，在四倍体水平上挖掘水稻的增产潜力成为水稻育种的新手段。同源四倍体水稻具有很强的遗传可塑性和很弱的遗传保守性，利用其作为水稻远缘杂交的桥梁，从野生物种中不断地引进有益的基因，这将有助于杂交水稻的多代利用和固定水稻的杂种优势。但是迄今为止，还没有关于同源四倍体水稻遗传多样性，遗传背景的报道。目前世界关于同源四倍体水稻的研究主要集中在中国，主要研究方向为培育、筛选结实正常的亲本材料，配置和筛选结实率正常或接近正常的组合。经过几十年研究，虽然在材料构建，细胞学研究等方面取得了较大进展，但同样由于结实率低的瓶颈问题未解决，而使多倍体水稻育种未能取得实质性进展。而近年来一些关于同源四倍体水稻低结实率机理的细胞学研究也由于缺乏统计学数据而缺乏说明性。本文用SSR标记，对选取的36个结实率正常同源四倍体水稻三系亲本和14个来源二倍体亲本，分析他们的遗传差异和群体遗传结构。本文还利用我们培育的高、低结实率的同源四倍体水稻恢复系、优良保持系和杂种F1及二倍体对照为材料，进行系统深入的细胞遗传学研究，进一步探讨同源四倍体水稻有性传递后代的发育过程，探索分裂期染色体行为特征与遗传性状稳定性的关系，为进一步选育多倍体水稻品种并将其应用于生产提供理论依据。同源四倍体水稻突变株D4063-1直链淀粉含量比来源二倍体明恢63下降一半,即其直链淀粉含量为5.23%，为研究其直链淀粉含量下降的原因，本文还根据普通水稻Wx基因设计引物，扩增测序获得了D4063-1Wx基因的全序列，与已报道Wx基因进行比对分析，并根据D4063-1和籼稻、粳稻的序列差异并根据D4063-1在该片段上的特征序列位点设计了用于识别D4063-1的寡核苷酸片段,为快速、准确的鉴别低直链淀粉的D4063-1创造了条件。 SSR标记具有基因组分布广泛、数量丰富、多态性高、容易检测、共显性、结果稳定可靠、实验重现性好、操作简单、经济、易于高通量分析等许多优点，被认为是用于遗传多样性、品种鉴定、物种的系统发育、亲缘关系及起源等研究的非常有效的分子标记。本研究选取了中国科学院成都生物所培育的同源四倍体和二倍体水稻亲本，并用36个微卫星标记进行了遗传差异和种群遗传结构分析。在50个品系中，我们观察到较高水平的多态性，每基因等位基因数（Ae）分布于2至6之间（平均值3.028），多态性信息含量（PIC）分布于0.04至0.76之间（平均值0.366）；期望杂合度（He）分布于0.04至0.76之间（平均值0.370），Shannon指数（I）分布于0.098至1.613之间（平均值0.649）。同源四倍体品系的等位基因数，期望杂合性和Shannon指数都比二倍体品系高。在供试50个品系中，较多材料均发现Rare基因，根据SSR多态性指数我们构建了同源四倍体和二倍体水稻的核心指纹库。F-统计值表明遗传差异主要存在于同源四倍体品系中（Fst=0.066）。聚类分析结果表明50个品系可以分为4个组。I组包括所有的同源四倍体和二倍体籼稻保持系，以及一个同源四倍体籼稻雄性不育系及其来源二倍体。II组仅包括IR来源的品系。III组比II组和IV组更复杂，包括同源四倍体和二倍体籼稻恢复系品系。IV组包括同源四倍体和二倍体粳稻品系。此外,由于等位基因及配子的遗传差异,同源四倍体与二倍体品系中存在单位点和双位点的遗传差异。分析结果表明,二倍体和四倍体水稻基因库的不同,其中遗传变异可以区分四倍体与二倍体水稻。同源四倍体水稻具有长期而独立的遗传性，我们能够选育并得到与二倍体亲本相比有特殊优良农艺性状的品系。 本研究以高结实率的同源四倍体水稻恢复系DTP-4、D明恢63及优良保持系D46B为材料进行农艺性状及细胞遗传学比较研究。DTP-4、D明恢63及保持系D46B的的染色体组成均为2N=4X=48，花粉母细胞具有较为理想的减数分裂行为，配对染色体的比率在99%以上，这与理论染色体组构成相符。DTP-4和D明恢63PMC减数分裂各个时期单价体和三价体的比例都非常低，而在MI， PMC观察到较多的二价体和四价体且四价体多以环状形式出现，其最大频率的染色体构型分别为12II 6IV和10II 7IV。恢复系DTP-4和D明恢63在MI四价体频率分别为2.00/PMC和2.26/PMC，而保持系D46B在MI四价体频率为6.00/PMC，极显著地高于恢复系品系，表明保持系D46B具有更好的染色体配对性质；AI保持系D46B的染色体滞后频率为10.62%，远低于恢复系材料DTP－4和D明恢63的19.44％和23.14％，接近二倍体对照明恢63的7.30%水平；TI保持系D46B具有比恢复系更低频率的微核数。而在TII，D46B的正常四分小孢子比率不但高于恢复系品系甚至高于二倍体对照。对高低结实率的同源四倍体水稻恢复系和杂种F1代的花粉育性，结实率和细胞遗传学行为进行了比较研究。DTP-4, D明恢63, D46A´DTP-4和D46A´D明恢63的花粉育性和结实率比D什香和D46A´D什香显著提高。减数分裂分析的结果表明，DTP-4，D明恢63，D什香，D46A´DTP-4，D46A´D明恢63和D46A´D什香其减数分裂染色体构型分别为：0.05I +19.96 II (9.89棒状+10.07环状) +0.01III + 2.20 IV, 0.11I +19.17 II (8.90 棒状+10.37 环状) +0.09III + 2.26 IV + 0.01 VI, 1.33I +9.46 II (4.50 棒状+4.96 环状) +0.44III + 6.02 IV + 0.09VI + 0.09 VIII, 0.02I +14.36 II (6.44 棒状+7.91 环状) +0.01III + 4.80IV + 0.01VIII, 0.06 I +17.67 II (11.01 棒状+6.67 环状) +0.06 III + 3.10 IV + 0.01 VI and 1.11 I +11.31 II (5.80 棒状+5.51 环状) +0.41 III + 5.63 IV+0.03VI+0.03VIII。在同源四倍体水稻恢复系和杂种F1代材料中，最常见的染色体构型为16II +4IV和12II +6IV。在减数分裂过程中，结实率较高的材料染色体异常现象较少而结实率较低的材料染色体异常现象较严重。在杂种F1代中，二价体的比例要低于其相应的恢复系亲本，同样的，单价体，三价体和多价体的比例相比其恢复系亲本也偏低。然而，在减数分裂MI，杂种F1代中四价体的比例要显著高于其恢复系亲本。在中期I，每细胞单价体的比例和花粉育性呈现出极高的负相关(-0.996)，当单价体数目升高时，花粉育性下降。其次是每细胞三价体的比例(-0.987)，之后则是每细胞多价体的比例与花粉育性的负相关(-0.948)。但是统计分析表明，二价体和四价体的比例对花粉育性和结实率没有显著影响。这一结果表明出了花粉育性和细胞减数分裂行为的相关性，同源四倍体的减数分裂行为为筛选高结实率的同源四倍体种系提供了理论依据。 突变体是遗传学研究的基本材料。利用突变体克隆水稻基因,并进而研究基因的生物学功能是水稻功能基因组学的重要研究内容。本课题组在多年的四倍体水稻育种研究中已获得多个低直链淀粉含量突变体，其中一些突变体在直链淀粉含量下降的同时，胚乳外观也发生了显著改变，呈半透明或不透明。同源四倍体水稻突变株D4063-1直链淀粉含量比来源二倍体明恢63下降一半,即其直链淀粉含量为5.23%。为研究其直链淀粉含量下降的原因，我们根据普通水稻Wx基因设计引物，扩增测序获得了D4063-1Wx基因的全序列，与已报道Wx基因进行比对分析；同源四倍体水稻D4063-1Wx基因最显著变化为在外显子序列中发生了碱基缺失，导致移码突变，在第9外显子终止密码子提前出现。D4063-1Wx基因碱基位点的变化还导致了其序列上的酶切位点的变化,对常用限制性内切酶位点分析分析结果表明同源四倍体水稻相对于籼稻和粳稻多了2个sph1酶切位点，相对于粳稻减少了6个Acc1，增加了4个Xba1，1个Xho1，1个Pst1和1个Sal1酶切位点。聚类分析表明D4063-1Wx基因序列与籼稻亲源关系较近，由此推测D4063-1Wx基因来源于籼稻的Wxa基因型。另外，根据D4063-1Wx基因的碱基差异，我们推测D4063-1Wx基因外显子碱基变化导致的RNA加工障碍是其直链淀粉降低的主要原因，并可能与其米饭较软等品质相关。本文还根据D4063-1和籼稻、粳稻的序列差异并根据D4063-1在该片段上的特征序列位点设计了用于识别D4063-1的寡核苷酸片段,并作为PCR反应的引物命名为AUT4063-1,将该引物与我们设计的扩增普通籼稻、粳稻的Wx基因引物F5配合使用建立了识别D4063-1的显性和共显性两种检测方式的分子标记,为快速、准确的鉴别低直链淀粉的D4063-1创造了条件。 研究同源四倍体水稻基因组的遗传差异，探索同源四倍体水稻的遗传规律，研究分裂期染色体行为特征与遗传性状稳定性的关系,旨在揭示四倍体水稻中同源染色体配对能力的遗传差异，为进一步选育多倍体水稻品种并将其应用于生产提供理论依据。 Autotetraploid rice (2N=4X=48, AAAA) is a new germplasm developed from diploid rice (2N=2X=24, AA) through chromosomes doubling with colchicines and is an excellent resource for desirable resistance genes to the pathogens and high protein content. Therefore, heterosis utilization on polyploidy is becoming a new strategy in rice breeding. At present, the main research on autotetraploid rice centralizes in China. Breeding effort has been made to improve autotetraploid rice genetically, however, the progresses are limited due to higher degree of divergence between hybrid sterility and polygenic nature. But to date, almost nothing is reported about the genetic diversity, original and genetic background of autotetraploid rice. Despite several reports on cytological analysis of the mechanisms of low seed set in autotetraploid rice still the results are inconclusive due to lack the statistical evaluation. Therefore, the study on the mechanisms of low seed set in autotetraploid is a priority for rice breeding. Microsatellites or simple sequence repeats (SSRs) are the widely used marker for estimating genetic diversity in many species, including wild, weedy, and cultivated rice. In our research, genetic diversity and population genetic structure of autotetraploid and diploid populations collected from Chengdu Institute of Biology, Chinese Academy of Sciences were studied based on 36 microsatellite loci. For the total of 50 varieties, a moderate to high level of genetic diversity was observed at population levels with the number of alleles per locus (Ae) ranging from 2 to 6 (mean 3.028) and PIC ranging from 0.04 to 0.76 (mean 0.366). The expected heterozygosity (He) varied from 0.04 to 0.76 with the mean of 0.370 and Shannon’s index (I) ranging from 0.098 to 1.613 (mean 0.649). The autotetraploid populations showed a slightly higher level of effective alleles, the expected heterozygosity and Shannon’s index than that of diploid populations. Rare alleles were observed at most of the SSR loci in one or more of the 50 accessions and core fingerprint database of the autotetraploid and diploid rice was constructed. The F-statistics showed that genetic variability mainly existed among autotetraploid populations rather than among diploid populations (Fst=0.066). Cluster analysis of the 50 accessions showed four major groups. Group I contained all of the autotetraploid and diploid indica maintainer lines and a autotetraploid and its original diploid indica male sterile lines. Groups II contained only original of IR accessions. Group III was more diverse than either group II or IV and comprised of both autotetraploid and diploid indica restoring lines. Group IV included japonica cluster of the autotetraploid and diploid rices. Furthermore, genetic differences at the single-locus and two-locus levels, as well as components due to allelic and gametic differentiation, were revealed between autotetraploid and diploid varieties. This analysis indicated that the gene pools of diploid and autotetraploid rice are somewhat dissimilar, which made a variation that distinguishes autotetraploid from diploid rices. Using this variation, we can breed new autotetraploid varieties with some new important agricultural characters but the diploid rice has not. Cytogenetic characteristics in restorer lines DTP-4, DMinghui63 and maintainer line D46B of autotetraploid rices were studied. DTP-4, DMinghui63 and D46B showed the advantage of high seed set and biological yield. The meiotic chromosome behavior was slightly irregular in DTP-4, DMinghui63 and D46B. We observed less univalent, trivalent and multivalent at MI, but more bivalent and quadrivalent were observed. The most frequent chromosome configurations were 12II 6IVand 10II 7IV in restorer and maintainer lines, respectively. The quadrivalent frequency of DTP-4 and Dminghui63 at metaphase(MI) was respectively 2.00/PMC and 2.26/PMC. However that frequency of D46B was 6.00/PMC, which was greatly significantly higher than DTP-4 and Dminghui63. That indicates the maintainer D46B has better chromosome pairing capability in metaphase (MI). The frequency of lagging chromosomes of the maintainer D46B at anaphaseI (AI) was 10.62%, which was significantly lower than that of DTP-4(19.44%) and Dminghui63(23.14%) and nearly reaching the level of diploid CK(7.30%). In telophaseI (TI) maintainer D46B showed lower frequency of microkernel at TI and lower frequency of abnormal spores at telophaseII(TII). We also studied pollen fertility, seed set and cytogenetic characteristics of restorer lines and F1 hybrids of autotetraploid rice. DTP-4, DMinghui63, D46A´DTP-4 and D46A´DMinghui63 showed significantly higher pollen fertility and seed set than DShixiang and D46A´DShixiang. Pairing configurations in PMC of DTP-4, DMinghui63, DShixiang, D46A´DTP-4, D46A´DMinghui63 and D46A´DShixiang were 0.05 I+19.96 II (9.89 rod+10.07 ring)+0.01 III+2.20 IV, 0.11 I+19.17 II (8.90 rod+10.37 ring)+0.09 III+2.26 IV+0.01 VI, 1.33 I+9.46 II (4.50 rod+4.96 ring)+0.44 III+6.02 IV+0.09 VI+0.09 VIII, 0.02 I+14.36 II (6.44 rod+7.91 ring)+0.01 III+4.80 IV+0.01V III, 0.06 I+17.67 II (11.01 rod+6.67 ring)+0.06 III+3.10 IV+0.01 VI and 1.11 I+11.31 II (5.80 rod+5.51 ring)+0.41 III+5.63 IV+0.03 VI+0.03 VIII, respectively. Configuration 16 II+4 IV and 12 II+6 IV occurred in the highest frequency among the autotetraploid restorers and hybrids. Meiotic chromosome behaviors were less abnormal in the tetraploids with high seed set than those with low seed set. The hybrids had fewer frequencies of bivalents, univalents, trivalents and multivalents than the restorers, but higher frequency of quatrivalents than the restorers at MI. The frequency of univalents at M1 had the most impact on pollen fertility and seed set, i.e., pollen fertility decreased with the increase of univalents. The secondary impact factors were trivalents and multivalents, and bivalents and quatrivalents had no effect on pollen fertility and seed set. The correlative relationship between pollen fertility and cytogenetic behaviors could be utilized to improve seed set in autotetraploidy breeding. The amylose content of autotetraploid indica mutant Rice D4063-1 dropped by half than diploid Minghui 63, that is, its amylose content of 5.23%.The whole sequence of Waxy gene of D4063-1 is amplified and sequenced. And the discrepancy of bases is found comparing to the reported Waxy gene. The Waxy gene of autotetraploid Rice D4063-1 had a base deletion in exon sequence, which resulted frameshift mutation in exon 9 and termination codon occur early. The mutation of Wx also led to the change of some common restriction endonuclease sites. Results showed compared to indica and japonica, D4063-1 had two adding sph1 sites. Compared to japonica, D4063-1 had six decreasing Acc1, a adding Xho1, Pst1 and Sal1 restriction sites. Phylogeny analysis shows that the DNA sequence of Waxy gene of D4063-1 is closer to Indica, and we suppose that the Waxy gene of D4063-1 is origin from genotype Wxa. In addition, according to the base differences of Wx in D4063-1, we deduce that RNA processing obstacle led by base change of intron is the main cause to low the amylose content, and related to phenotype of its soft rice. Based on analysis of fragments of D4063-1, indica and japonica and according to the special point of the three species, primers as markers-AUT4063-I were designed for distinguishing the D4063-1 from other rice. Combining with primer pair F5, dominant and codominant ways were established for discriminating them., rapid and correct identification of D4063-1 from other rice could be done. The genetic analysis is important to ensure the original of autotetraploid rice, for maintaining the “distinctiveness” of autotetraploid varieties, and to differentiate between the various genetic background of autotetraploid rice. The autotetraploid breeding will benefit from detailed analysis of genetic diversity in the germplasm collections. Further investigation on mechanisms of meiotic stability should benefit polyploid breeding. These findings demonstrated opportunity to improve meiotic abnormalities as well as grain fertilities in autotetraploid rice.

糯小麦杂交后代的基因型鉴定及Wx基因的效应研究

糯小麦在食品加工业、淀粉加工业及其它工业上有着重要用途，是近年来许多国家小麦研究的重要课题。国外糯小麦选育尚未突破高产与糯性相结合的难点，国内目前还没有培育出高蛋白强筋型的糯小麦品种，这在一定程度上与缺乏合适的育种方法和高效、实用的糯小麦分子标记辅助育种技术有关。国内外对Wx基因效应的研究主要利用缺体－四体系、重组自交系或近等基因系，还未见有利用遗传背景相同的BC5F2代回交改良群体的报道。 糯性位点近等基因系是小麦淀粉品质育种的重要材料，而我国目前还没有一套中国栽培小麦遗传背景的糯性位点近等基因系。为了选育部分糯小麦、全糯小麦和中国栽培小麦遗传背景的糯性位点近等基因系，我们利用Wx蛋白电泳和高效实用的分子标记技术体系来鉴定糯小麦杂交后代的基因型，结果证明该体系能有效地用于糯小麦的分子标记辅助育种。以中国春糯性位点全套近等基因系为研究材料，对小麦Wx基因的6个STS标记和1个CAPS标记进行了筛选，改良PCR扩增条件以及产物检测方式后，从这些标记中筛选出3个标记，包括鉴定Wx-A1、Wx-D1位点的2个共显性STS标记和Wx-B1位点的1个显性STS标记。利用上述3个分子标记从BC5F2 代回交改良群体中筛选出了8种Wx基因型，经卡方检验，其分离比符合3对基因的分离比例，其中基因型为aabbdd的植株有2株，直链淀粉含量分别为1.81%和0.82%，为全糯小麦；基因型为AAbbdd，aabbDD的部分糯性植株各有1株，直链淀粉含量分别为15.24%和17.57%。以上4株植株的农艺性状和品质性状接近回交亲本“川育12”，并明显优于全糯材料“98Y1441”，表明采用回交法与Wx基因分子标记辅助选择相结合，有助于培育高产、优质的全糯和部分糯小麦。同时，本研究中建立的分子标记技术体系，也为选育具有中国栽培小麦遗传背景的糯性位点近等基因系奠定了基础。 在基因型鉴定的基础上，利用糯小麦杂交后代BC5F2代回交改良群体研究了各基因缺失降低直链淀粉含量的效果和各基因合成直链淀粉的能力，以及直链淀粉含量与农艺性状、品质性状、淀粉糊化特性等的相关性。缺失不同Wx基因的8种基因型，直链淀粉含量差异显著。研究单缺失基因型发现，减少效应最大的是Wx-B1b基因，Wx-B1b和Wx-D1b基因没有显著差异，减少效应最小的是Wx-A1b基因。研究双缺失基因型发现，Wx基因合成直链淀粉的能力，Wx-B1a基因最高，Wx-A1a基因最低，而Wx-B1a和Wx-D1a基因差异很小；直链淀粉含量与株高、穗长、小穗数、穗粒数、千粒重等农艺性状相关不显著，表明淀粉品质育种可以与高产育种实现有机结合；直链淀粉含量与SDS-沉降值呈显著负相关(r=-0.726)，说明直链淀粉含量降低在一定程度上有利于提高小麦营养与加工品质，这一结果至今未见有文献报道；全糯类型的淀粉糊化特性与其他类型显著不同，具有最高的峰值粘度和稀懈值，最低的低谷粘度、最终粘度、反弹值、峰值时间、糊化温度、起始糊化温度，表明糯小麦淀粉在食品和工业上具有特殊用途；稀懈值与直链淀粉含量呈极显著负相关(r=-0.969)，其他粘度参数与直链淀粉含量呈显著正相关(最终粘度r=0.797，低谷粘度r=0.910、反弹值r=0.954、峰值时间r=0.970、糊化温度r=0.962、起始糊化温度r=0.932)。以上结论可为糯小麦品种选育和淀粉品质改良提供理论依据。 Waxy wheat is very important in food processing industry, starch processing industry and the other industries, so it is a focus of wheat research in many countries these years. Foreign wheat breeders have not conquered the difficulty of high yield combined with waxy character, and there is no waxy wheat variety with high protein and strong gluten in China at present，all of which were caused by lacking proper breeding methods and effective, applied molecular markers-assisted selection technique at a certain extent. Until now, research about the effect of waxy genes mainly depended on nullisomic-tetrasomic lines, recombinant lines or near-isogenic lines, and it is lacking the reports of using improved BC5F2 backcross progenies under the common genetic background. Near-isogenic lines at the Wx loci are important materials for wheat starch quality breeding. However, there are no such lines under the Chinese cultivated wheat genetic background. To develop partial waxy wheats, waxy wheats and near-isogenic lines at the Wx loci under the Chinese cultivated wheat genetic background, we use SDS-PAGE of waxy proteins and effective, applied molecular marker-assisted selection technical system to identify the genotype of waxy wheat’s progenies. The results indicated that such a system is applicable in waxy wheat’s molecular marker-assisted selection effectively. A series of Chinese Spring Wx loci near-isogenic lines were used to identify the specific bands of six STS markers and one CAPS marker of Wx genes. After optimizing PCR amplification and separating of PCR products, three co-dominant and dominant STS-markers were identified at the Wx-A1, Wx-D1 and Wx-B1 loci, respectively, which were used to identify the genotype of waxy wheat’s progenies. Eight Wx genotypes were developed from the improved BC5F2 backcross progenies, which follows Mendelian segregation. Among them, there were two aabbdd waxy plants whose amylose content in the BC5F3 seeds were 1.81% and 0.82%, respectively. In addition, there were partial waxy plants (AAbbdd and aabbDD) whose amylose content in the BC5F3 seeds were 15.24 % and 17.57%, respectively. Most agronomic and quality traits of these four plants resembled those of the recurrent parent “Chuanyu 12”, and superior to waxy wheat parent “98Y1441”. This shows that backcross approach in combination with molecular marker-assisted selection of waxy genes is helpful to develop partial and full waxy wheat with good traits in the waxy wheat breeding program. At the same time, molecular marker-assisted selection technical system in this paper, also establish the base for breeding the near-isogenic lines at the Wx loci under the Chinese cultivated wheat genetic background. According to the results of genotype identification, we use waxy wheat’s improved BC5F2 backcross progenies to verify the effects of null alleles on reducing amylose content and determine the amylose synthesis capacity of each Wx gene independently, and investigate the relativity among amylose content with agonomic traits, quality traits, starch pasting properties respectively. There was significant difference in amylose content of the eight genotypes carrying different null Wx alleles. The reducing effect of the single null alleles was the most significant in Wx-B1b, followed by Wx-D1b and Wx-A1b, and there was no significant difference between Wx-B1b and Wx-D1b. The results of the double null lines further demonstrated that for the capacity of amylose synthesis, Wx-B1a was the highest, followed by Wx-D1a and Wx-A1a, and there was no significant difference between Wx-B1a and Wx-D1a. Amylose contents of the eight genotypes were not significantly correlated with plant height, spike length, spikelets per spike, grains per spike, 1000-grain weight, which showed that starch quality breeding could integrate with high yield breeding. Amylose contents of the eight genotypes were negatively significantly correlated with SDS-sedimentation value(r=-0.726), which suggested that reduction in amylose content is propitious to improve quality at a certain extent. Starch pasting properties of the full waxy type was significantly different from the other seven types, with the highest peak viscosity and breakdown, and lowest valley viscosity, final viscosity, setback, peak time, pasting temperature and starting pasting temperature. It indicated that waxy wheat starch has special use in food and industry. Breakdown was negatively significantly correlated with amylose contents (r=-0.969), and the other parameters were positively significantly correlated with amylose contents (r=0.797 for final viscosity, r=0.910 for trough viscosity, r =0.954 for setback, r =0.970 for peak time, r=0.962 for pasting temperature and r=0.932 for starting pasting temperature ). The results of the study are very useful for waxy wheat variety breeding and starch quality improvement.