大二上遗传学ch 7 chromosome mapping in eukaryotes

1、Chapter 7Linkage and Chromosome Mapping in Eukaryotes 连锁遗传分析(真核生物的连锁遗传与染色体作图）1500 genes7.1 Linkage & INDEPENT ASSORTMENTobserve deviations from9 : 3 : 3 : 1 ratios derived from dihybrid crosses双因子1 : 1 : 1 : 1 ratios derived from test crosses测交 1906 sweet pea cross： Bateson and Punnett found phenome

2、non with non independent assortment ：Cross 1、Ppurple, long pollenXred, round F1purple, long pollen Expected9331cross 2P11rple a11drou11cl (PPII) X reel a11cl l011g (ppLL)p紫花、) 1圆花粉粒 x 红花、长花粉PP 1tppLL1氖花、长花趴沪P1h卫PpLlF,紫、长1红、长红、圆总_ L.ppL_ppll、五个体数22s119511 1971 1t.t 19235.8 1 178.5 11 78.5Ii 26.2419箕的

3、埋论Coupling phase and repulsion phaseCoupling phase: Allelic arrangement in,vhich mutant are on the same chromosome and ,vild-t)rp e alleles areon the homologue.Repulsion phase: allelic arrangement in,vhich each homologous chromosome has mutant and , Tild- t ) rp ea ll eles.声 coupling phaseversusre11

4、ulsion 11hase二描述两个显性性状连在一起遗传两个隐性性状连在一起遗传= I圆花粉粒（隐）一个显性性状与另一个隐性性状匡霆I圆花粉粒（隐）一个隐性性状与另一个显性性状= I长 花粉粒（显）Independentassortment is true for:Genes on separate chromosomes.Genes on the same chromosome (linked genes) if they are far apart. This phenomena were first explained in 1911 by two Drosophila genetici

5、sts, Thomas H. Morgan and his undergraduate student, AlfredH. Sturtevant.A simplified overview of the major theme of this chapter is given in Figure 7-1, which contrasts the meiotic consequences of (a) independent assortment, In Figure 7-l(a)Independent assortment (genes on different chromosomes or

6、very far apart on one chromosome):abFreq.AaBbParentalaabbParentalAabbRecombaaBbRecombABab Ab aBlinked genes, same chromosomeswrite as . Aa Bb or AB/ab or Ab/aBA Ba bcis orcouplingorA ba Btrans orrepulsion2 possible dihybrids, illustrate as .segregate and assort dependently distancetest cross to ab/a

7、b 4 progeny types .in CIS: 0 Ab : 0 aB : 0 AB : 1 Ab : 0 abcompare these results with what occurs if the same genes are linked on the same chromosome. If no crossing over occurs between the two genes Figure 7- 1 (b), only two genetically different gametes are formed.Each gamete receives the alleles

8、present on one homolog or the other, which transmits intact as the result of segregation. This case demonstrates complete linkage, which produces only parental or noncrossover gametesAB ab ababComplete linkage (genes too close on the chromosome for crossing-over to occur):abFreq.AaBbParentalaabbPare

9、ntalAabbRecomb0aaBbRecomb0ABab Ab aBexchange generates two new allele combinations, calledrecombinant or crossover gametes.-hX,p害“I- 1富b厅巨。蠡B,晶bAB ”:谝， B咋AB， 曼 l -，！ 厂l： 汒= .圈 -.1 .L，唱：IAS-,aba b-3，一: 1 1AB ab ababIncomplete linkage:AaBbParentalaabbParentalAabbRecombaaBbRecombabRatioABab Ab aBMore t

10、han 50% of offspring with parental arrangement of genes and fewer than 50% with alleles recombined.7.2 Incomplete Linkage,Crossing over, and Chromosome Mapping It is highly improbable that two randomly selected genes linked on the same chromosome will be so close to one another along the same chromo

11、some that they demonstrate complete linkage. Instead, crosses involving two such genes almost always produce a percentage of offspring resulting from recombinant gametes. This percentage is variable and depends on the distance between the two genes along the chromosome. This phenomenon was first exp

12、lained in 1911 by two Drosophila geneticists, Thomas H. Morgan and his undergraduate student, Alfred H. Sturtevant.1=Recombinant valueSingle CrossoversWhy should the relative distance between two loci influence theamount of recombination and crossing over observed betweenthem?During meiosis, a limit

13、ed number of crossover events occurs in each tetrad. These recombinant events occur randomlyalong the length of the tetrad. Therefore, the closer two loci reside along the axis of the chromosome, the less likely any single crossover event will occur between them. When a single crossover occurs betwe

14、en two nonsister chromatids, the other two chromatids of the tetrad are not involved in this exchange and enter the gamete unchanged. Even if asingle crossover occurs 100 percent of the time between two linked genes, recombination is subsequently observed in only 50 percent of the potential gametes

15、formed. This concept is diagrammed in Figure 8-6.1 Two-point testcross两点测验Recombination AnalysisRecombination analysis is a technique used to determine how frequently a crossover occurs between two genes during meiosis and, therefore, how far apart the genes are on the chromosome.To do recombination

16、 analysis, you need:1) A heterozygote for two genes known to be on the same chromosome.2) A homozygous recessive to test cross it to (so that every genotype will have a unique phenotype).3) Enough offspring for accurate counts of non- crossover and crossover progeny.What is the distance between two

17、X-linked genes in Drosophila?w = white eyes;w+ = red eyesm = miniature wings;m+ = normal wings Inbred stock #1white eyes, miniature wingsInbred stock #2wmwmwmYred eyes,w + m +w + m +normal wings (wildtype)w +m+Y1) Make an F1 hybrid2) Testcross it.w + m + wmw + m + wmwmYTestcross:w + m + wmwmYGametes

18、:w m or YF2 Phenotypes:Pw+ m+w+ m+/ (w m or Y)red, normal376Pw mw m/ (w m or Y)white, miniature367Rw + mw+ m/ (w m or Y)red, miniature217Rw m+w m+/ (w m or Y)white, normal230Total1,190Calculate the percentage of offspring that resultfrom crossing over = 217 + 230 / 1190 x 100.Recombinant Frequency (

19、RF) number ofrecombinan ts 100total progeny 447 100= 37.6%1190 white eyeminiature wingsX chromosome37.6% recombinant frequency (RF)37.6 map units37.6 centiMorgans (cM)Another mapping experiment:pr = purple eyes,pr+ = red eyesvg = vestigial wings, vg+ = normal wingsInbred stock #1 pr + vg +(Wildtype)

20、pr + vg +Inbred stock #2prvgprvg(Purple eyes, vestigial wings)1) Make an F1 hybrid2) Testcross it.List the gametes each testcross parent will produce:pr + vg +prvg pr + vg + prvg(Wildtype phenotype) prvg prvg pr + vg + prvgprvgprvgpr vgPpr+ vg+pr+ vg+/ pr vgred, normal165Ppr vgpr vg/ pr vgpurple, ve

21、stigial175Rpr+ vgpr+ vg/ pr vgred, vestigial20Rpr vg+pr vg+/ pr vgpurple, normal22Total382Total CO frequency (RF) = 11.5% (20 + 22 /382 x 100)purple eyevestigial wings11.5% recombinant frequency11.5 map units11.5 centiMorgans (cM) 图距两个基因在染色体图上距离的数量单位称为图距（map distance）。 l重组值（交换值）去掉其百分率的数值定义为一个图距单位（ma

22、p unit， mu），后人为了纪念现代遗传学的奠基人THMorgan，将图距单位称为“厘摩（centimorgan，cM），1cM1重组值去掉的数值。例 表3-6果蝇的一些性连锁基因的重组频率基因重组频率（重组值）(y)和白眼(w)0.010(y)和辰砂眼(v)0.322(y)和小翅(m)0.355辰砂眼(c)和小翅(m)0.030白眼(w)和辰砂眼(v)0.300白眼(w)和小翅(m)0.327白眼(w)和斜截翅(r)0.450辰砂眼(v)和斜截翅(r)0.2690 1.0 32.2 35.5 58y w vm rmechanism of crossing over?1. Linkageg

23、enes on the same chromosome show up a certain amount of recombinants 换言之，在两对基因连锁遗传时，形成亲代所没有的新组合的机制 是什么？ Janssens（1909）（chiasmatype hypothesis）：（1） at miosis, diplonemapaired chrs criss-cross forming chiasma . Thisis the site at which exchange occurs between homologous chr（2） linked genesexchangingea

24、ch otherresult in recombination between non alleles图3 l 1配对的同濮染色体有两个交叉交叉 结 处表 示 非姊 妹染色 单体 间发 生过 交换交叉绪L ep to n e m a C hrom os om 令S are al 戊 ady d1.11plicated- S ynapUnemal complex begin s 伈 ap pear_Zyg o n e m a藏 Pairing记initia ted . Syn 叩 tin em a.l co m ple x develops more fu ll y.IP a ci h y;n

25、e m a Pa il1i1n.g i令co mp !et 啄i .嘈 C hrom os om es thi ck en .- Cmssin.9 over occurs. Chmmosorne bouquot 伈rmsD叩lome.ma Repulsfon between homo iogs, beg in s. C h:iasm ata ara clear1y vis1hle .- Cho m o的 mo sa,re he l d t o g 呻 e r at chiasmata and contmmeroC 匈平均tit i 郢17 , 切，心 n” nr的 汹 文， n仁 :AII r

26、ig tl饬 亿 眨 lr.,.，配 ；一D ia ki n e, s i 巴 M ax imum,chro m oso 江th ic ke ni ngO OOUr s. .Ch r.asma.t已l d i 租 ppea r. Ghrnmo isom es move to己 qua mr na l pkane.Sister Chromatid Exchanges Knowing that crossing over occurs between synapsed ho-mologs in meiosis, we might ask whether such a physical exchan

27、ge occurs between homologs during mitosis. While homologous chromosomes do not usually pair up or synapse in somatic cells (Drosophila is an exception), each individual chromosome in prophase and metaphase of mitosis consists of two identical sister chromatids, joined at a common centromere. Surpris

28、ingly, several experimental approaches have demonstrated that reciprocal exchanges similar to crossing over occur between sister chromatids.Three traditionalinheritance laws三大遗传定律的关系 segregation , independent assortment are the basis oflinkagelaws 自由组合定律和连锁交换定律是变异的主要机制 自由组合是染色体间重组（分配）(interchromosom

29、al recombination) 交换则是染色体内重组(intrachromosomal recombiation)连锁群通过连锁遗传研究，得出两条基本规则：（1） 如果A基因与B基因连锁，B与C连锁，A与C也连锁（2） 如果A基因与B基因连锁，B与C不连锁，A与C也不连锁。基因位于染色体上，那么同一条染色体的不同基因就组成 了一个连锁群。理论上讲，每种生物有多少对染色体，就 有多少个连锁群。基因定位的常用方法 两点测验 twopoint testcross 三点测验 threepoint testcross两点测验 具有2对相对性状的个体杂交 F1与隐性个体测交 根据测交结果判断2对基因是

30、否连锁 如果连锁，重组率是多少例 玉米籽粒：有色C无色c ，饱满（Sh）凹陷sh，非糯性Wx对糯性（wx）。 为了明确这三对基因是否连锁，曾有人做过三个两点测验。第1个测验有色饱满CCShSh ccshsh无色凹陷有色饱满 CcShsh ccshsh无色凹陷Rf13.6%即3.6cM第2个测验饱满糯性 wxwxShSh WxWxshsh凹陷非糯饱满非糯WxwxShsh wxwxshsh凹陷糯性Rf2 20% 即20cM2个两点测验结果1：C-c和Sh-sh3.6%2：Wx-wx和Sh-sh20%第3个测验非糯有色 WxWxCC wxwxcc糯性无色非糯有色WxwxCc wxwxcc糯性无色Rf

31、3 23.6%即23.6cM有了第3个测验结果，就能确定3个基因座的相对位置了。两点测验的局限性： 工作量大，烦。 当遗传距离大于5cM，准确性不高。 三点测验与基因定位 Morgan1911曾提出，重组值的大小可能由基因在染色体上的距离决定。这个设想可以用实验的方法来验证. 例如a.b.c三个基因相互连锁，可用两两间a-b、b-c、a- c间的重组值确立三者之间的关系。但这样做很烦琐。 Morgan与他的学生Sturtevant提出了一个包括三个基 因的一次 配测定三者关系的方法。既三杂合体abc/+或ab+/+c,与三隐性个体abc/abc测交。这种测验称三点测验。一次测验等于三次两点测验

32、。其优点： （1）一次测验得到三个重组值，在同一基因型背景同 一环境。 （2）可得到两点测验得不到的数据双交换值。 看一下黑腹果蝇中三点测验的例子：Three-point testcrossRecombination occur at Calculation of each recombinant phenotype observed percentTotal number例2:有双交换的情况: ec ct +/+ + cv ec ct cv/y表型个体数目比例交换发生在ecct+2125+cv2207ec+cv273+ct+265ec+217ec-ct ec-cv ct-cv81.5%-10

33、.1%+-eccvct+ctcv2238.3%+-+10.28.4+518.4+0.2ecctcv30.1%-+合计5318100% 18.4%10.2%8.4%ec-cv=10.2%ct-cv=8.4%ec-ct=18.4%，而+=18.6%这是什么原因呢？看一下+和ec.ct.cv8只果蝇ltC:(;Cl-一士一-十 一-一-一-一-一-一-一- ecct-J- -,- , -+ec,e rlec+ -， ct+.-.-.zee+ctL L , ,+C V十图-6J.,5只 有 当C V 位 千 中 问 时 ，同 时 发 生 两 个 单 交 换 后才可以产生双交换个体和吐从上图可以看出，e

34、c-ct之间发生了两次交换。只有当cv/+存在时，才能识别出双交换的发生。所以在计算ec-ct交换值时，一定要加上2倍的双交换值：ec-ct的交换值=18.4 + 2 X 0.1%=18.6%。这个法则叫做基因直线排列定律。三点测验的总结：（1） 三杂合体的创造（2） 三杂合体与三隐性测交（3） 每两个基因为一组统计重组合的比例（4） 直线定律排列基因顺序（5） 根据6种或是8种表型，8种表型中，个体数最少 的表型为双交换产物，计算双交换值。Multiple CrosseoverIt is possible that in a single tetrad, two, three, or mor

35、e exchanges will occur between nonsister chromatids as a result of several crossover events. Double exchanges of genetic material result from double crossovers (DCO), as shown in Figure 7-7. For Supposethatcrossovergametesresultingfrom single exchanges are recovered 20 percent of the time (p = 0.20)

36、 between A and Band 30 percent of the time (p = 0.30) between B and C. The probability of recovering a double-crossover gamete arising from two exchanges, between A and B and between B and C, is predicted to be (0.20)(0.30) = 0.06, or 6 percent. It is apparent from this calculation that the frequenc

37、y of double- crossover gametes is always expected to be much lower than that of either single-crossover class of gametes.可见每发生一次单交换,对其邻近的交换产生影响, 它的邻近也发生一次交换的机会要减少一些,这种现象称 为干涉(干扰).一般用并发系数或并发率（coincidence）来表示干扰的 大小，公式如下：并发系数(coefficient of coincidence)C=观测到的双交换率/两个单交换率的乘积I=1-C上例: C=0.15%/(10.2%*8.4%)=

38、0.17 I=1-C=0.87 I=1-C If interference is complete and not double crossovers occur, then I=1.0; If fewer DCOs than expected occur, I is a positive than number and positive interference has occurred; If more DCOs than expected occur, I is a negative than number and negative interference has occurred;

39、Positive interference is most often observed in eukaryotic systems.遗传干涉和并发率1 遗传干涉的概念2 并发率 = 实际双交换/理论双交换= 实际双交换/单交换I X 单交换II= 0 (干涉完全)干涉=1-并发率并发率01 (部分干涉) (10) = 1 ( 无 干 涉 )正干涉(postive interference)负干涉(negaive interference)7.4The Accuracy of Mapping Experiments 两个基因座在染色体上距离较远，其间可能发生双交换或四交换等更高数目的even-

40、 numbered exchanges (偶数)次交换，形成的配子却仍然是非重组型的，在这种情况下， 若简单地把重组率看作交换率，交换率就要被低估，图距自然也随之缩小了。显然随着重组率的增大，其衡量图距的准确性也随之下降。(1.5+.06)+(4.0+.06）=5.62A Mapping Problem in Maize Instead of using the gene symbols and superscripts (e.g., bm+ ,v+, and pr+), we simply use + to denote each wild-type allele. This system

41、is easier to manipulate, but requires a better understanding of mapping procedures In maize, the recessive mutant genes bm (brown midrib), v (virescent 淡 绿 seedling), and pr (purple aleurone 糊 粉 层 ) are linked on chromosome 5. Assume that a female plant is known to be heterozygous for all three trai

42、ts. We do not know: (1) the arrangement of the mutant alleles on the maternal and paternal homologs of this heterozygote, (2) the sequence of genes, or (3) the map distances between the genes.i(b) Actual results of maip ping ,cro ss.IPhe not ype.s of offspringNumberTotal and percentage46742志1%16122.

43、314.5%+7.8395IExch ang, e classificationIN!o ncrossover (NCO)S11 n1(i1I e cir心 so ver (SCO)Sin 1 le c1ros soverVbm2:30pr十|237十bm82prV|79V|200pr+bm195prVbm44十十|4235 .6%+7.843.4(SCO)867 置8%1Duubl.e crossover (DCO)气比罕 盓的tI心？品; ,IEplanation谓vndbmpr住）Noncrossover pheni0lyp巳p心 vii.de eb, asi 5(b)pr十l、七Ibm

44、 ndof de归 m in1a1in固g ttie correct a rrange,m ent ore.s. on homo1.ogs压 pec ted double l:re 邸 刃 er phenoty伪 5忏 V is 5llil the middleVpr十十-tVprand bmVprbm(,c)bm vpr(d) v压pected dollJble G 0匈 侂 r phenolyp缸if bm is inthe middl仑Expec也d double u o笠0四 叮 phe notypesand十十计p r i5 in me midd,le (This 1s t he-

45、哎 tuoisituoti O几），(e) V加VprG泗 n trhoad（ou, tca)and (d) amcorrocL 5inglepl(nv+bmandbmV+cn5$Ove ptb p叭heeniot ype5 when exchange occu sen va1 nd pr十p,and”订bmG 泗othpOrCado(”ut“act妇)a,rpitwhe(dedenn)oty妞pa卢红n0釭呴d 位如e nI,en由xgchl.e( 91) VFinal mapprbmSignificanceofa GeneticMapA genetic map tells us:(1) W

46、hich genes are on a particular chromosome(2) The order in which the genes occur on the chromosome(3) An indication of the relative (but not absolute) distances between genes.abc某一染色体上三个隐性基因连锁图为0103，0(1) 如果完全干涉, (2)如果无干涉, (3)如果干涉系数40%，(4)如果并发系数(符合系数)50%, 在abc/+abc/abc杂交中预期表型频率为多少？连锁图和连锁群u 位于同一对同源染色体上

47、的所有基因座组成一个连锁群（linkage group），它们具有连锁遗传的关系。u 把连锁基因之间的顺序和距离标志出来， 就成为连锁图（linkage map），又称为遗传学图（genetic map）果蝇的连锁图7.6 Somatic Cell Hybridization and Human Gene Mapping体细胞杂交定位原理与技术 Cell culture ：1907 Hannision first neuron Cell clone：1956 Puck and Marcus single mammalian cellto proliferate a clone in vitro

48、 Mutant cell culture:培养突变细胞系体系的建立，运用体外单细胞培养技术培养和克隆特定的突变型细胞。 Somatic Cell fusionHybridization Splenocyte fusion with myeloma cells to screen positive hybridomas to produce monoclonal antibody Researches showed，fusion can be made between various animal cells . rat、Cricetidae 中国仓鼠、rabbit and human fibr

49、oblast 、macrophage 、lymphocyte、leukocyte etc.两亲 细胞融合过程可分两个阶段： 异核阶段：细胞核有来自本的核，随后异核体进行有丝分裂，核膜，组合在一起形成一个细胞核的杂种细胞。 杂种细胞形成阶段：由于两种细胞的不同，细胞核将发生不同的变化。 Cells of two different species are fused the heterokaryon 异 核 阶 段 initially has 2n chromosomes from both cells. After the nuclei fuse, there is a tendency to randomly lose chromosomes during subsequent mitotic cell divisions. Result A series of clones (杂种细胞形成 )with different combinations of the original double set of chromosomes.Somatic -cell Hybridization 例如：小鼠 X 中