13 第13章 分子标记辅助选择.pptx

第1页：分子标记辅助选择目录

• 分子标记辅助选择
• 分子标记辅助选择的基本原理
• 质量性状的标记辅助选择
• 数量性状的标记辅助选择
• 分子标记辅助选择的挑战与

第2页：传统的育种主要是根据植株的表现型进行选择

传统的育种主要是根据植株的表现型进行选择，而环境条件、基因间互作、基因型与环境互作等多种因素都会影响表型选择效率

第3页：抗逆性、抗病虫性

F2P2F1P1xlarge populations consisting ofthousands of plantsPHENOTYPIC SELECTIONField trialsGlasshouse trialsDonorRecipient传统育种Salinity screening inphytotro Bacterial blight screeningPhosphorus deficiency plot

第4页：育种者在实践中探索运用遗传标记来提高育种

• 育种者在实践中探索运用遗传标记来提高育种的选择效率与育种预见性
• 遗传标记包括形态学标记、细胞学标记、生化标记与分子标记
• 以DNA多态性为基础的分子标记，在作物遗传图谱构建、重要农艺性状基因的标记定位、种质资源的遗传多样性分析与品种指纹图谱及纯度鉴定等方面得到广泛应用，尤其是分子标记辅助选择（molecular marker-assisted selection，MAS）育种更受到人们的重视

第5页：种质资源、作物产量

• Molecular plant breeding expands useful geneticdiversity for crop improvement.
• Molecular plant breeding increases favorable geneaction.
• Molecular plant breeding increases the efficiency ofselection.Molecular plant breedingMoose S P,& Mumm R H. Molecular plant breeding as the foundation for21st Century crop improvement. Plant Physiology,2008,147:969-977

第6页：育种与选择、优缺点与发展策略

Breeding technology pipeline from past to present to future

第7页：建立分子设计育种体系种质资源重测序数字表

建立分子设计育种体系种质资源重测序数字表达谱进化基因组学关联分析连锁分析突变体库转基因关键基因50-100个基因组图谱高效能新品种种质资源简易基因芯片选择效率提高1000倍育种周期缩短到1/2-2/3

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第9页：分子标记与基因位点、作物产量

• Molecular plant breeding
• Marker-assisted selection
• Genomic selection
• Genetic engineering
• Design breedingMark Tester* and Peter Langridge. Breeding Technologies to IncreaseCrop Production in a Changing World.Science327, 818 (2010)

第10页：概念：通过基因定位找到与目标基因紧密连锁

• 1、概念：通过基因定位找到与目标基因紧密连锁的分子标记后，可通过该分子标记间接地对目标性状进行选择
• 此法称分子标记辅助选择(Molecular Assistant Selection, MAS )
• MAS是育种中的一个诱人领域,将给传统的育种研究带来革命性的变化
• MAS主要应用在有利基因的转移和基因的累加等方面
• Tester M & Langridge Breeding technologies to increase cropproduction in a changing world. Science,2010,V 327:818-822

第11页：RR(1-r)20.9025抗性供体

RR(1-r)20.9025抗性供体受体×RS2r(1-r)0.095SSr20.0025目的基因与标记连锁（交换值为r）亲本中的标记带型F1中的标记带型F2群体中3种标记带型当r=0.05时，根据标记基因型mm选择目的基因型RR，选错的概率约为0.10共显性DNA标记的辅助选择原理×mRMSmRMS×

第12页：分子标记与基因位点、倍性与染色体

Definitions1)Phenotypic selection (PS)–based on phenotypic value2)Marker-based selection (MBS)–from markers that represent QTL or are linked to QTL3)Marker-assisted selection (MAS)–from a combination of phenotypic value and marker information4)Marker-assisted backcrossing (MABC)5)Marker-assisted recurrent selection (MARS)6)Genomic Selection or Genome-wide Selection (GS orGWS)–Select for breeding values summed across many markers withoutestimation of QTL

第13页：倍性与染色体、环境与污染

• (1) selection without test crossing or a progeny test;
• (2) selection independent of environments;
• (3) selection without laborious fieldwork orintensive laboratory work;
• (4) selection at an earlier breeding stage;
• (5) selection for multiple genes and/or multipletraits;
• (6) whole genome selection

第14页：抗病虫性、倍性与染色体

• Useful if conventional screening methods are laborious,costly, or environmentally dependent–Selections for disease and insect resistance can be made in theabsence of the pathogen or pest–Greatest potential advantage over phenotypic selection for traitswith low penetrance or low heritability
• May reduce population sizes needed for phenotypicselection–May permit selection of individual plants
• May speed up the breeding process–May be effective for early generation testing–Selections at the seedling stage can be a great advantage in cropswith a long generation time–Reduce number of generations in a backcrossing program byselecting for recovery of the recurrent parent genome as well asgenes of interest from the donor parent2、Most suitable for MAS

第15页：分子标记与基因位点、抗病虫性

• Pyramid genes for a single trait that could nototherwise be distinguished at the phenotypic level–Accumulating multiple quantitative trait loci (QTL) fordisease resistance may provide a higher level ofresistance and/or more durable resistance to changes inthe pathogen population–MAS may promote deployment of fewer resistancegenes
• Genes for multiple traits of interest may also becombined in one cultivar with relative efficiency
• Marker technologies provide the potential tounderstand the underlying causes of epistasis andGXE, which could greatly improve selectionefficiency

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第17页：环境与污染、育种与选择

• Qualitative traits and quantitative traits with highheritabilityare more amenable to MAS than quantitativetraits with low heritability, which is generally the case forphenotypic selection as well.
• Epistasis (or effect of genetic background) andgenotype by environment interactions (GXE)canconfound progress from MAS just as they do inconventional selection schemes.
• Efforts to improve the precision of QTL estimationthrough increased replication and multilocational testingwill also increase the efficiency of phenotypic selection,thereby reducing the gains that may be attained throughMAS.
• Catch 22:–If phenotypes are poor indicators of genotypes, you cannot mapQTL for use in MAS–If phenotypic data are good, you don’t need MASParadox of MAS

第18页：分子标记与基因位点、分子遗传

1)How tightly it is linked to genes controlling importanttraits.2) The relative importance of those genes in determiningthe phenotype.3)The consistency of linkage disequilibrium between themarker and QTL4) The frequency of the QTL (MAS will be more beneficialwhen the QTL is in low frequency)3.The utility of a marker depends on

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第20页：分子标记与基因位点、育种与选择

• Requirements for wide-scale application of MAS
• Validation of QTL in breeding materials–Multiple markers in vicinity of QTL desirable
• Simple, quick, inexpensive protocols for tissuesampling, DNA extraction, genotyping, and datacollection
• Efficient data tracking, management, andintegration with phenotypic data
• Decision support tools for breeders–optimal design of selection strategies–accurate selection of genotypes

第21页：大多数情况下

• 大多数情况下，质量性状无需借助于分子标记，但采用分子标记辅助选择可提高选择效率：○表现型测定难度大或费用太高
• ○表现型只能在个体发育后期才能测量，而育种实践中希望在早期选择
• ○除目标性状外，还需对遗传背景进行选择
• ○质量-数量性状的选择

第22页：标记辅助选择的基本方法

• 1、标记辅助选择的基本方法1）前景选择（foreground selection）——对目标基因的选择
• 单标记可靠性：取决于标记与目标基因间连锁的紧密程度。如，标记座位M/m与目标基因座位Q/q连锁，重组率为r，则在F2根据标记基因型M/M获得目标基因型Q/Q的概率为p=（1-r）2。选择正确率随重组率增加而迅速降低

第23页：分子标记与基因位点、人口增长与粮食安全

• ★如要求正确率在90%以上，则标记与目标基因间的重组率必须＜5%
• Genotypes of F2plants forXa-21gene as determined by a PCR marker and progeny testsAccuracy（%）F3progency testPCR analysisNo. of plantsGenotypeNo. of plantsGenotype91.231RR34RR3Rr85.74RR28Rr24RrThe distance between marker pTA248 andXa-21gene isabout 1cM (Ronld and Tanksley, 1991).The 9% inaccuracymay reflect the error in phenotyping or the variation inrecombination frequency from mapping population to breedingpopulation.

第24页：如要求选到：株目标基因型的概率为P

• ★如要求选到
• 1株目标基因型的概率为P，则必须选择带有目标基因型M/M的植株的最少数目为：n=log（1-P）2/log（1-r）2即使重组率高达
• 0.3，也只要选择
• 7株具有基因型M/M的植株，就有99%的把握能保证其中有
• 1株为目标基因型；而如果不用标记辅助选择（相当于标记与目标基因间无连锁，即r=0.5），则至少需要

第25页：双边标记可靠性将大大提高

• 双边标记可靠性将大大提高
• 如标记M1/m1和M2/m2各位于目标基因座Q/q的一侧，与目标基因间的重组率分别为r1和r2，则：F1的基因型为M1QM2/m1qm2F1产生的标记基因型为M1M2的配子有两种类型，包含目标等位基因的M1QM2和包含非目标等位基因的双交换M1qM2，由于双交换发生频率很低，因此，在后代中通过同时跟踪M1和M2来选择目标等位基因Q，正确率必然很高

第26页：在无干扰时，F2代通过选择标记基因型M1

• 在无干扰时，F2代通过选择标记基因型M1M2/ M1M2而获得目标基因型Q/Q的概率为：p=（1-r1）2（1-r2）2/[（1-r1）2（1-r2）+r1r2]※在两标记间的图距固定的情况下，r1= r2时p最小（即目标基因位于两标记间的中点）
• ※在实际情况下，单交换间一般总是存在相互干扰的，双交换的概率更小，因此双标记选择的正确率比理论期望值更高

第27页：分子标记与基因位点、人口增长与粮食安全

Marker-assisted identification ofPi-z5gene in a segregating population with flanking markersProgeny testPredictio Accuracy ofpredictionrrRrRRRG64RG456100%25(25)aRR(31)aRR426Rr44RR97.70%43144Rr53Rr5rr1Rr100%2424rr25rra: Number of plants predicted to carry the indicated genotype.The distance between RG456 and RG64 is about 14.4cM.

第28页：背景选择：参考完整的分子遗传图谱

• 2）背景选择（background selection）———对除了目标基因之外的其他部分（遗传背景）的选择在F2，根据两个相邻标记，可推测出它们之间的染色体区段的来源和组成
• 参考完整的分子遗传图谱，检测所有标记基因型，最后绘制出图示基因型（graphic genotype）

第29页：12222222222212222321

• 122222222222122223211222222222212222333312333322111A品种×B品种的F2代一个个体的图示基因型，共
• 5对染色体，白色表示来自A品种的区段，黑色表示来自B品种的区段，红色表示发生了交换的区段；竖杠表示标记所在位置。22

第30页：标记辅助选择在育种上的应用

• 2、标记辅助选择在育种上的应用
• 1）多基因聚合（gene pyramiding）——将分散在不同品种中的有用基因聚合到一个基因组中

第31页：分子遗传（分子标记辅助选择）

A gene-pyramidingscheme cumulating six target genes.（Servin等，2004）

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第33页：个抗稻瘟病基因在染色体上的位置例子1

3个抗稻瘟病基因在染色体上的位置例子1：通过分子标记辅助选择方法聚合水稻抗稻瘟病基因（Zhengel al. 1995）

第34页：C101LAC×C101A51C10

C101LAC×C101A51C101LAC×C101PKTF1F1××F2150株F2150株用相邻标记进行选择Pi-1Pi-2Pi-1Pi-410个Pi-1和Pi-2纯合的植株10个Pi-1和Pi-4纯合的植株××F2150株标记辅助选择F13个抗病基因纯合的植株三个近等基因系

第35页：分子标记与基因位点、抗病虫性

例2：N. Huanget al（1997）Pyramiding of bacterial blight resistance genes in rice:marker-assisted selection using RFLP and PCRDNA marker-assisted selection was used topyramid four bacterialblight resistance genes,Xa-4,xa-5,xa-13andXa-21. Breeding lines withtwo, three and four resistance genes were developed and tested forresistance to the bacterial blight pathogen (Xanthomonas oryzaepv.oryzae).The pyramid lines showeda wider spectrum and a higher levelof resistancethan lines with only a single gene.

第36页：近等基因系接种株数标记分析株数

近等基因系接种株数标记分析株数

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第40页：分子标记辅助回交育种

• 2）分子标记辅助回交育种（gene transfer，MAB）将供体亲本中的有用基因转移或渗入到受体亲本的遗传背景中，从而达到改良受体亲本个别性状的目的
• 在每一代都需通过对供体等位基因的选择使目标基因座位保持杂合，同时提高目标座位以外的其他染色体区域的受体基因组（Recipient genomecontent，RGC）比例

第41页：分子标记与基因位点、倍性与染色体

• Marker-assisted backcrossing
• Foreground selection：markers can be used incombination with or to replace screening for the targetgene or QTL.
• Recombinant selection:markers can be used to selectBC progeny with the target gene and recombinationevents between the target locus and linked flankingmarkers.
• Background selection:Markers unlinked to the targetlocus can be used to select BC progeny with the greatestproportion of recurrent parent genome.

第42页：前景选择：通过标记基因型的选择来控制目的

• (1)前景选择（正选择）保证从每一回交世代选出的作为下一轮回交亲本的个体都包含目的基因
• 通过标记基因型的选择来控制目的座位基因型，可通过TCR值来评定效果

第43页：Target

Target Control Rate (TCR)：对任一回交后代个体，当其标记基因型为杂合的“供/受”时，其目标座位基因型也为“供/受”的概率。换句话说，一个个体在标记为期望基因型，而目标座位不是期望基因型的概率为（100-TCR）。Target Control Rate = TCR(%)= Pr { (TD/TR) | (MD/MR) }×100= Pr { (TDMD/TRMR) } / Pr { (MD/MR) }×100TD和TR为目标座位上供体和受体的等位基因；MD和MR为标记座位上供体和受体的等位基因；Pr{(X/Y)}表示基因型X/Y的概率，|表示条件

第44页：分子标记与基因位点、分子遗传

It is clearly seen thatcontrol of the target by a single marker isnot satisfactory in most cases. The marker must be as close as 1 cMto the target to keep the risk of ‘losing’ the target below 5% overfive BC generations.Even with a single marker at 1 cM, the risk oflosing the target is close to 10% in BC10. For greater distances of asingle marker, the risk becomes rapidly too high.Target controlled by single marker（Hospital，2003）单标记

第45页：当目标座位为两侧标记界定时

• 当目标座位为两侧标记界定时，即使标记相距较远，效果也是很好的，如两侧标记分别相距10cM时，其控制效果与单标记距离在1cM的效果相近
• 很显然，这种结果的原因在于当有两侧标记时，要打破标记与目标的连锁需要进行双交换
• 而单标记情况下，只需要发生单交换
• Target controlled bytwo flanking markers（Hospital，2003）双标记

第46页：回交：回交

• 回交
• 1回交
• 2回交
• 3回交
• 6传统回交育种
• 75.0 87.7 93.399.0标记辅助
• 85.5 98.0100.0回交育种回交育种中轮回基因组百分比Young and Tanksley，1989，计算机模拟
• (2)背景选择（负选择）▲加快遗传背景恢复成轮回亲本基因组的速度，以缩短育种年限

第47页：指有利基因和不利基因间的连锁

• 指有利基因和不利基因间的连锁，使回交育种在导入有利基因的同时也带入了不利基因，常常造成性状改良后的新品种与原目标不一致.传统的回交育种中，无法鉴别目标基因附近所发生的遗传重组，只能靠碰巧来选择消除了连锁累赘的个体
• 大致需要100代才能基本消除连锁累赘的目的
• ▲避免或减轻连锁累赘(linkage drag)的问题

第48页：用高密度的分子标记连锁图就有可能直接选择

• 用高密度的分子标记连锁图就有可能直接选择到在目标基因附近发生了遗传重组的个体
• 只要在BC1和BC2中进行标记辅助选择，即可得到含有目标基因的供体染色体片段长度不大于2cM的植株，从而只需两个回交世代就可达到基本消除连锁累赘的目的

第49页：传统回交育种标记辅助回交育种年F1BC1

• 传统回交育种标记辅助回交育种年F1BC1BC2BC3BC20BC100年
• 0.511.5黑色代表供体基因组；白色代表轮回基因组（Tanksleyet al. 1989）0.511.5210.550.5

第50页：分子标记与基因位点、倍性与染色体

Efficiency of a typical marker-assistedbackcross scheme（Hospital，2003）Noselectio RGC%Homozygosityat selected markers(%)populatio Generatio Non-carrierchromosomeCarrierchromosome757960.638.470BC187.592.287.473.6100BC293.79898.893150BC396.999100100300BC4Target-marker distance on the carrier chromosome was 2 cM. Eachnon-carrier chromosome was controlled by three markers.

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第52页：分子标记与基因位点、育种与选择

• Example: Marker
• assisted transfer ofXa21: strategyMinghui 63×IRBB 21(Xa21)Minghui 63×F1Minghui 63×BC1F1Minghui 63×BC2F1BC3F1BC3F2(Xa21)Selection for recombination oneither side of theXa21locusSelection for recombination on theother side of theXa21locusSelection for Minghui 63 backgroundMinghui 63 (Xa21)
• Selection forXa21homozygotesChen ShengZhang Qifa

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第54页：分子标记与基因位点（分子标记辅助选择）

MH63(Xa21)MH63(CK)

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第56页：分子标记与基因位点、倍性与染色体

• 4.2.3Introgression of genes from wild-germplasm
• The transfer of few genes or small segments ofchromosomes in interspecific crosses is verytedious on account of unwanted transfer ofunadapted gene complexes from wild species.
• Such small segments, however, can be selectivelytransferred with the help of molecular markerslocated on the specific segment of chromosome ofwild species to minimize linkage drag.

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第60页：123456 7注

• 1为桂夏豆
• 2号；2，3，4，5，6，7为桂夏豆
• 2号×（桂夏豆
• 2号×Z4）

第61页：回交与基因渗入、种质资源

Introgression of genes from wild-germplasm

第62页：受体亲本

• 受体亲本（华粳籼74）
• 供体亲本（26个品种）
• 受体亲本
• 分子标记选择代换片段BC Fn受体亲本：代换系文库：图中的细线条表示基因组，粗线条表示来自供体的代换片段。华粳籼74单片段代换系文库的建立水稻单片段代换系文库的构建

第63页：数量性状的标记辅助选择意义很大

• 数量性状的标记辅助选择意义很大，但困难也很大，目前多局限在理论研究上，实际成功例子很少
• （1）只有少数数量性状的全部QTL被精细定位出来，无法对数量性状进行全面的标记辅助选择
• （2）育种过程中同时对许多目标QTL进行选择相当困难
• （3）上位性效应对选择效果影响很大，常使选择达不到预期目的

第64页：标记值选择直接依据个体的加性效应值进行选

• 1、标记值选择直接依据个体的加性效应值进行选择，能够提高选择效率
• 如何估计出各个个体的加性效应值？用性状—标记回归的方法来计算出个体的标记值（marker score），并以该值作为个体加性效应值的估计

第65页：首先筛选出对目标性状效应明显的标记★然后

• ★首先筛选出对目标性状效应明显的标记★然后根据它们的加性效应估计值计算出个体的标记值：m= ∑aixi+ε（ai为第i标记的加性效应值，Xi：第i个标记的基因型指示变量，对应于基因型MM、Mm、mm分别取
• 0、-1；n为筛选的标记数，ε为环境机误）以个体标记值为依据的选择——标记值选择

第66页：标记型值选择与表型值选择之间的相对效率R

• 标记型值选择与表型值选择之间的相对效率RMMP=△GM/△GP=√p/h2△GM和△GP分别为标记型值选择与表型值选择的遗传进度，p为标记所能解释的加性遗传方差（σM2）占总的加性遗传方差（σG2）的比例，h2为狭义遗传力对遗传力高的性状，表型值选择效率已经很高，无须采用标记选择
• 对遗传力较低的性状，标记型值选择具有较高的相对效率，而且遗传力越低，其相对效率就越高

第67页：基因型选择：基因型选择

2、基因型选择

2、基因型选择

（1）基因型选择

即对每个目标QTL利用其两侧相邻标记或单个紧密连锁的标记进行选择，其原理和方法与质量性状相似

（2）关键问题

The uncertainty of the truetarget location，i.e. the exact location of thetarget is often not known

第68页：计算TCR值

• （3）计算TCR值：假定QTL检测已经完成，QTL的期望（最可能）位置已经确定，同时已给出置信区间，而真实的目标是位于期望QTL附近的某一点，遵循一个均值为
• 0、方差为置信区间长度的正态分布。对每一个可能的真实位置都计算TCR值，然后平均（Hospital and Charcosset，1997）

第69页：不完全基因型选择

• （4）不完全基因型选择，即对那些已定位的QTL进行基因型选择
• 在选择目标QTL的同时，同样也可以利用分子标记进行背景选择，加快遗传背景恢复成轮回亲本的速度

第70页：分子标记在水稻产量遗传改良上的应用通过分

• △分子标记在水稻产量遗传改良上的应用通过分子技术在野生稻中，发现两个增产QTL，每个QTL具有比对照品种增产18%的效应（Xiao等，1995）。已将其中的一个QTL转移到栽培稻中，用这种携带野生稻增产QTL水稻配组的杂交稻，2003年在浏阳作双季晚稻百亩示范，亩产700公斤
• 3、QTL的回交转移

第71页：

第72页：J23A/Q611

J23A/Q611

第73页：分子标记与基因位点、育种与选择

Shen et al. (2005) Used BNL1521 and BNL2961 markers forQTLfs21 selection, and BNL1122 for QTLfs22 selection. Thefiber strengthof the selected individual plants showed the greatimprovement.△分子标记在棉花纤维强度遗传改良上的应用

第74页：抗逆性、分子遗传

M. Wissuwa á M. Yano á N. Ae,Mapping of QTLs for phosphorus-deficiency tolerance in rice (Oryza sativa L.) TheorAppl Genet (1998) 97: 777Ð783△A success story in MAS on rice Pi efficiency

第75页：分子标记与基因位点、抗逆性

• M. Wissuwa
• J. Wegner
• N. Ae
• M. Yano,Substitution mapping of Pup1: a major QTL increasing phosphorus uptake of ricefrom a phosphorus-deficient soil.Theor Appl Genet (2002) 105:890–897

第76页：倍性与染色体（分子标记辅助选择）

Graphical genotypes for the Kasalath segmenton chromosome 12 of 160 selected F3individuals of six F2families, as well as the meansand standard deviationsfor tiller numbers ofgenotypic classes within families

第77页：分子标记与基因位点、倍性与染色体

Sequence comparison ofphosphorusuptake1(Pup1) loci and flanking regions in three ricevarieties. A Nucmer alignment of the unmasked Kasalath sequence assembled from three BACclones with the corresponding sequences in genotypes Nipponbare (japonicareference genome)and 93-11 (indicareference genome) is illustrated.Sigrid Heuer, et al.Comparative sequence analyses of the major quantitative trait locusPhosphorusuptake1(Pup1) reveal a complexgenetic structure.Plant Biotechnology Journal(2009)7, pp. 456–471

第78页：分子标记与基因位点、分子遗传

Chin JH, et al., Development and application of gene-based markers for themajor rice QTL Phosphorus uptake 1. Theor Appl Genet (2010) 120:1073–1086

第79页：分子标记与基因位点、分子遗传

Chin JH, et al., Development and application of gene-based markers for themajor rice QTL Phosphorus uptake 1. Theor Appl Genet (2010) 120:1073–1086Pup1 haplotype of 58 rice genotypes. The Pup1 haplotype was determined bygenomic PCR using seven Pup1 gene-based markers (K41–K59) and two closelyflanking markers (RM28073, Ba76H14_7154).

第80页：分子标记与基因位点、种质资源

Pup1 haplotype survey. Rice accessions genotyped with the Pup1 markerswere grouped according to their preferred cropping systems and varietal group(indica, japonica, aus) (a). In (b) the same genotypes were grouped accordingto their classification as modern or traditional varieties.Chin JH, et al., Development and application of gene-based markers for themajor rice QTL Phosphorus uptake 1. Theor Appl Genet (2010) 120:1073–1086

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第82页：分子标记与基因位点、抗逆性

Root elongation and reduction in shoot length under P deficiency in hydroponics culturesolution. A set of contrasting Pup1 near-isogenic lines with the Pup1 locus (NILC443,NIL6-4, and NIL14-4) and sister lines without the Pup1 locus (NIL6-3, NIL14-6) weregrown in hydropnics solution with 0 and 100 lM P, respectively

第83页：分子标记与基因位点、抗逆性

Chin JH, et al., Developing Rice with High Yield under PhosphorusDeficiency: Pup1 Sequence to Application. Plant Physiology, 2011, Vol. 156,pp. 1202–1216Pup1 gene models and positions of markers.

第84页：分子标记与基因位点（分子标记辅助选择）

Sequence comparisons of marker amplicons

第85页：分子标记与基因位点、分子遗传

PCR amplicons of Pup1 gene-specific markers in representative rice varieties.

第86页：分子标记与基因位点（分子标记辅助选择）

Pup1 haplotype in diverse rice genotypes

第87页：分子标记与基因位点（分子标记辅助选择）

Pup1 core markers

第88页：分子标记与基因位点、育种与选择

Field evaluation and selection of IR64-Pup1 and IR74-Pup1 breeding lines

第89页：育种与选择、优缺点与发展策略

1. ChallengesFrom a breeder’s view, MAS must not only beefficient, but more efficient than alreadyavailable breeding methods on an economical .第四节分子标记辅助选择的挑战与发展策略

第90页：抗病虫性、作物产量

For clear-cut cases(i) provides an increased genetic gain per unit oftime, which is the case when MAS permitsselection at DNA level at an early stage in thedevelopment and/or when it permits reductionof the generation interval (most trees andanimals, crops with breeding based on progenytesting),(ii) provides an increased genetic gain per unit ofcost (molecular genotyping cheaper thanphenotypic evaluation, disease resistances,…).

第91页：抗病虫性、作物产量

• For less clear-cut cases
• the success of MAS experiments (in plants) appears todepend mostly on the complexity of the geneticarchitecture of the trait.
• MAS must surely be more efficient when the trait is simple(e.g., resistance) as opposed to complex (e.g., yield),controlled by few QTLs (as opposed to many QTLs), whenthe QTL effects are large (not small), when there are no G9 E interactions, and in general when there are no epistaticinteractions (although a counter example was given above)

第92页：育种与选择、分子遗传

1) Finding new sources of major genes(i). the apparently huge potential of plant geneticresources to provide new genes of agronomicinterest.(ii). to integrate metabolic, phenotypic and genomicdatabases to allow a wider view of the plantmetabolome and the application of this knowledgewithin genomics-assisted breeding .

第93页：分子标记与基因位点、环境与污染

• 2) Accelerated methods of gene pyramiding
• If large sources of major genes were really to be unlocked,as wishfully anticipated in the previous section, then anefficient marker-assisted gene pyramiding scheme wouldneed to tackle multiple, possibly linked, genes, frommultiple parents.3).Managing and controlling QTL stability
• The (in-)stability of QTL effects (across environments,genetic background…), even for QTLs with reasonablylarge effects, is one of the main causes of unexpected resultsof MAS experiments.
• Assessing the effects of each QTL in each environment is animmense task.

第94页：育种与选择（分子标记辅助选择）

• 4). Dealing with polygenic complex traits
• Some traits have been turned from ‘complex’, quantitative,traits into ‘simple’Mendelian traits after discovering that theywere controlled by a surprisingly low number of QTLs withlarge effects.
• To optimize the model selection process and find a compromisebetween the power of detection, the risk of false positives andof over fitting the predictive model.5).Integrating MAS in traditional breedingpractices
• to incorporateMAS in existing breeding schemes, in order toprovide additional gain at limited cost.
• the use of MAS would certainly be facilitated if it was betterintegrated into traditional breeding practices, rather thanconsidered as a substitute.

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第96页：分子标记与基因位点、诱变与突变体筛选

2.1 Factors to moderate the acceptance of MAS（1）The markers developed need to be relevantto the selection criteria of the breeding program.(2) The extent to which the breeder can readilyphenotypically select for a characteristic, withoutthe more costly use of markers (Morris et al.2003).(3) Even for some of the more complex traits,there can be a lower-cost screening method thanthe application of a molecular marker.2. Economic assessment of MAS

第97页：分子标记与基因位点、抗病虫性

(4) The variation in the extent to which markersare applicable with different breeding populations,as validation can be time-consuming andexpensive.(5) When marker correlation with characteristicsidentified by phenotypic screening is incomplete,breeders face uncertainty as to the extent towhich they can rely on the marker rather than thephenotypic screening.(6) Although markers for particular resistance (orlack thereof) against pests or diseases can bevaluable, the ever-evolving nature of suchdiseases means that other phenotypic evaluationwill still be needed.

第98页：分子标记与基因位点、育种与选择

(7).The extent to which molecular biologists areworking closely with the breeders towardscommongoals is important if marker developmentisto be coordinated with the needs of breeders.(8).Reliable markers have not yet been developedformany characteristics of interest to breeders.(9).The initial linkage between the marker and thedesirable trait may break down, requiring thebreeder to periodically conduct phenotypicassessment of the breeding lines to ensure thatMAS remains relevant and useful.

第99页：分子标记与基因位点、品质性状

• 2.Economic assessment of MAS2.2 Costs of marker-assisted selection1).the cost and conditions of the intellectual property relating to that marker;
• 2).the costs of tissue collection and storage;
• 3).the method of extraction of DNA/protein for analysis;
• 4).the extent to which extracted DNA is used for a number of different markers;
• 5).the analytical method used;
• 6).the rate of throughput of the analytical system;
• 7).the degree to which the analysis utilizes multiplexing and multi-pooling;
• 8).the ease of scoring the alleles after the analysis;
• 9).the unit costs of the labor for the different operations (Dreher et al. 2003);
• 10).the capital cost of the laboratory equipment required; and11).the utilization of the equipment required (that is, the overall throughput of thelaboratory)

第100页：分子标记与基因位点、分子遗传

2.Economic assessment of MASComparison of costs of molecular marker with phenotypic evaluations, hypotheticalAustralian program

第101页：分子标记与基因位点、种质资源

• Improving the efficiency of MAS
• Better precision and sharing of phenotypic data
• Simultaneous development of multiple mapping populations forparticular traits, using a set of common parents
• Selection for haplotypes
• Techniques for continual readjustment of QTL estimates overcycles of selection have been developed (“mapping as you go”)(Podlich et al., 2004)
• Association mapping techniques–more robust measures of QTL across diverse germplasm–more precise QTL mapping
• BLUP to account for information from relatives
• Better definition of QTL x environment interactions–specify breeding objectives for well-defined target environments

第102页：倍性与染色体、育种与选择

3.Stratigies1）A greater level of integration among conventionalbreeding, QTL mapping/validation and MAS.Genome resequencingIntegration of omics dataset

第103页：

第104页：回交与基因渗入、杂交与授粉

（2）Careful planning and execution of QTLmapping studies (especially for complexquantitative traits) and an emphasis on validatingresults prior to MAS.Advanced backcross QTL analysisAssociation mapping

第105页：

第106页：回交一代定位

• ◆回交一代定位，然后利用定位结果指导各回交世代的个体选择
• ◆回交高代QTL分析（advanced backcross QTLanalysis），可建立起一套受体亲本的近等基因系（NILs）或染色体片段代换系（CSSLs，SSSLs），从而对有关QTL进行精细定位，根据这一定位结果可大大提高标记辅助选择的可靠性
• QTL定位与选择

第107页：分子标记与基因位点（分子标记辅助选择）

（3）optimization of methods used in MAS such asDNA extraction and marker genotyping, especiallyin terms of cost reduction and efficiency.

第108页：分子标记与基因位点、倍性与染色体

• (4)Genome-wide selection for QTL
• Initially asked how markers could enhance traditionalselection in breeding programs
• Now it may be feasible to ask how best to use markers tomake genetic gains
• discovery of SNPs and other high density polymorphicmarkers
• high throughput genotyping has reduced costs
• costs for phenotyping continue to increase
• May be possible to select an array of markers withoutestablishing significant associations with traits
• Genomic selection (=genome-wide selection) firstproposed by Meuwissenet al., 2001Bernardo and Yu, 2007

第109页：分子标记与基因位点、倍性与染色体

Genome-wide selection1.In atraining population(both genotypic andphenotypic data available), fit a large number ofmarkers as random effects in a linear model toestimate all genetic effects simultaneously for aquantitative trait. The aim is to capture all of theadditive genetic variance due to alleles with bothlarge and small effects on the trait.2.In a breeding population (only genotypic dataavailable), use estimates of marker effects to predictbreeding values and select individuals with the bestGEBVs (genomic estimated breeding values).

第110页：分子标记与基因位点、倍性与染色体

• Genome-wide selection simulation study
• Simulation studies of testcross performance ofdouble haploids in maize showed genome-wideselection consistently more effective thanmarker-assisted recurrent selection (MARS)
• Greatest advantage for complex traits controlledby many QTL with low heritability
• Minimum number of markers = 128 to 256
• Most beneficial for recurrent selection (lesseffective for choosing parents of breedingpopulations or selection of single-crosses)Bernardo and Yu, 2007

第111页：分子标记与基因位点、育种与选择

Genomic selection will lead to improvedbreeding strategies that use largeamounts of genomic information, pairedwith estimated breeding values assignedto markers ⁄ haplotypes to expedite thebreeding process and increase the rate ofgain.4.4 Genomic selection

第112页：育种与选择、分子遗传

Genomic selection scheme

第113页：育种与选择、分子遗传

Schemes of genomic selection (GS) (left) and traditional MAS forthe selection of quantitative traits (right).

第114页：分子标记与基因位点、作物产量

• ShuYJet al.Genomic selection of seed weight based on low-densitySCAR markers insoybean.Genet Mol Res.2013 Jul 3;12(3):2178-88.
• Genomic selection (GS) was introduced into crop breeding toachieve more accurate predictions by considering all genes ormarkers simultaneously. We used dozens of sequence-characterized amplified region (SCAR) markers for genotypingsoybean varieties, and we identified markers associated withhundred-seed weight. The best linear unbiased predictor andBayesian liner regression methods were used to construct GSmodels to predict the hundred-seed weight trait based upongenotype information for trait selection. Both GS models showedgood prediction performance in soybean, as the correlationcoefficient between genomic estimated breeding values and truebreeding values was as high as 0.904. This indicated that GS wasperformed effectively based on dozens of SCAR markers insoybean; these markers were of low density but easily detectable.Therefore, the combination of GS modeling and highly effectivemolecular marker technology involving SCAR markers canfacilitate genetic breeding in soybean.

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第116页：分子标记与基因位点、育种与选择

• The differences between the MAS and GS
• (1) in the training phase, quantitative trait loci(QTLs) are identified in traditional MAS whileformulae for GEBV prediction are generated in GS,known as GS models;
• (2) in the breeding phase, genotype data are onlyrequired for targeted regions in traditional MAS,whereas GW genotype data are considered to benecessary in GS;
• (3) in the breeding phase, favourable individuals areselected based on the genotypes of markers in MAS,whereas GEBVs are used for selection in GS.

第117页：育种与选择、分子遗传

Nakaya A et al, Will genomic selection be a practical method for plantbreeding? Annals of Botany 110: 1303–1316, 2012

第118页：人口增长与粮食安全、育种与选择

• Genomic Selection Methods
• Relatively new approach–many questions remain–Composition of training population–Selection criteria–Choice of statistical models
• Stepwise regression, Ridge regression, Bayesian
• See review by Heffneret al., 2009
• Prospects for plants are bright (more efficient and lesscostly than conventional MAS)
• Results to-date are largely from simulations
• More empirical results should be available soo Hamblin, Buckler, and Jannink. 2011. Trends in Genetics 27: 98-106.

第119页：育种与选择、分子遗传

6 Breeding by design万建明作物分子设计育种。作物学报，2006，32：455-462Peleman J D and Van der Voort J R. Breeding by design.Trends Plant Sci,2003,8:330-334 作物分子设计育种将在多层次水平上研究植物体所有成分的网络互作行为和在生长发育过程中对环境反应的动力学行为;继而使用各种“组学”数据,在计算机平台上对植物体的生长、发育和对外界反应行为进行预测;然后根据具体育种目标,构建品种设计的蓝图;最终结合育种实践培育出符合设计要求的农作物新品种。Based on understanding the genetic basis of all agronomicallyimportant characters and the allelic variation at those loci, thebreeder would be able to design superior genotypes ‘in silico’

第120页：分子设计育种三个步骤

• 分子设计育种三个步骤
• Mapping loci involved in all agronomically relevanttraits（基因定位）
• Assessment of the allelic variation at those loci（等位基因鉴定）
• Breeding by Design（设计育种）△设计优良基因型△收集包括目的基因在内的各种生物分子信息△设计出可能方案组合，从中选择出最佳实施方案组合△采用转基因技术、MAS、基因敲除、RNA干涉和常规育种相结合的育种技术体系，实现目标基因的有机重组

第121页：分子标记与基因位点、倍性与染色体

Example of an introgression line (IL) library construction process in tomato(12 chromosomes each separated by a grey bar) using Marker-AssistedBackCrossing

第122页：分子标记与基因位点、倍性与染色体

Chromosome haplotypes. Chromosome haplotypes of a set of maizelines for one chromosome

第123页：分子标记与基因位点、回交与基因渗入

The principle of Breeding by Design. Subsequent selfings (F2) and BackCross(BC) selections using markers lead to the desired superior elite line genotype

第124页：抗逆性（分子标记辅助选择）

Overall model of an abiotic stress responseExample

第125页：抗逆性、作物产量

Design of stress-tolerant crops

第126页：分子标记与基因位点、作物产量

• 参考文献
• 1.Collard B C Y, Mackill DJ. Marker assisted selection: anapproach for precision plant breeding in the twenty-first century.Phil.Trans.R.Soc.B,2007,1-162.Bernarardo R. Molecular and selection for complex traits in plants:learning from the last 20 years.Crop Science, 2008,48:1649-1664

第127页：讨论：MAS的基本流程和优势

• 讨论
• MAS的基本流程和优势？
• MAS对哪些性状的选择有效？
• 如何提高数量性状的MAS选择效率？
• MAS广泛应用于育种实践中存在的问题？
• 如何在育种计划中结合MAS？