(完整版)chapter6thebacteriaandvirusinheritance

1、第六章 细菌和病毒的遗传escherichia colit2 phage第一节 细菌和病毒遗传研究的意义一、细菌基因组（以大肠杆菌为例）：dna主要以单个主染色体(main chromosome) 的形式存在，长约1100m，分子量约为2.6 109；不与组蛋白相结合；质粒(plasmid)plasmids: smaller circlesof dna that do not carryessential genes. techniques of cultivationall organisms need an energy source, a carbon source,nitrogen,

2、sulfur, phosphorus, several metallic ions, and water. heterotroph that require an organic form of carbon .autotroph that can utilize carbon as carbon dioxide.auxotroph（营养缺陷型）prototroph (wild type)原养型（野生型）a culture medium that has only the minimal necessities required by the bacterial species being g

3、rown is referedto as minimal medium.colony (clone)（克隆）平皿分离和摇瓶培养平皿分离和摇瓶培养bacterial phenotypes colony morphologycolony morphology relates simply to the form, color, and size of the colony that grows from a single cell. nutritional requirementsthe phenotype reflects the failure of one or more enzymes i

4、n the biosynthetic pathways of the bacteria.anabolic functional mutantscatabolic functional mutants e.g a strain of bacteria that can the sugar galactose as an energy source would be gal+. if it could not utilize galactose it would be termed gal. resistance and sensitivitythe phenotype involves resi

5、stance and sensitivity to drugs, phages, and other environmental insults.e.g. penicillin, an antibiotic that prevents the final stage of cell-wall construction in bacteria, will kill growing bacterial cells. penspenr二、病毒三、细菌和病毒在遗传研究中的优越性 世代周期短 易于管理和进行化学分析 便于研究基因的突变 便于研究基因的作用 便于基因重组的研究 便于用作研究基因的结构、功能

6、及调控机制的材料 便于进行遗传操作第二节第二节 噬菌体的遗传分析噬菌体的遗传分析一、噬菌体的结构（一）烈性噬菌体( virulent phage )animation(二) 温和性噬菌体( temperate phage )temperate phageanimation溶原性( lysogeny )：噬菌体侵入后，细菌并不裂解，噬菌体dna随细菌dna的复制而复制。原噬菌体( prophage )：温和性噬菌体dna整合入细菌dna上后，称为原噬菌体。二、t2噬菌体的基因重组与作图bacteriophage phenotypes plaque morphology. growth chara

7、cteristics on different bacterial strains. (host range)（宿主范围）r形成小噬菌斑，且边缘模糊。r形成大噬菌斑，且边缘清晰。h能侵染大肠杆菌菌株b，但不能侵染大肠杆菌菌株b/2。h能侵染菌株b和菌株b/2。h rh+r+hr hr+ h+r h+r+表型基因型透明，小半透明，大半透明，小透明，大hrhrhrh r%100总噬菌斑数总噬菌斑数重组噬菌斑数重组值hrrhrxh rxh，出现的4种噬菌斑数目和求得的重组值 0.959.039.00.7rchrh6.456.032.05.9rbhrh12.042.034.012.0rahrhrhrh

8、rhrh重组值每一基因型的%杂交%0 .2410024%3 .121003 .12%6 . 11006 . 1作连锁图rbrcrahrahrcrbrahrbrcrahrcrbrcrb rcrbrcrb的重组值rbh=12.3 排列顺序是rc-h-rbt2 噬菌体的连锁图也是环状的。三、 噬菌体的基因重组与作图类型数目占总数重组频率(%)亲本类型 975s co1 mi924单交换型s 302.9% co1 mi32单交换型s co1 615.3% mi51双交换型s mi50.86% co1 13合计20913.76 6.16 8.2 噬菌体s co1 mi 的杂交结果及分析sco1mi3.7

9、66.168.220.86第三节 细菌的遗传分析actually, bacteria have four different methods to gain access to foreign genetic material : transformation, conjugation, sexduction, and transduction. 一、transformation(转化)transformation was first observed in 1928 by f. griffith and later (in 1944) examined at the molecular le

10、vel by o. avery and his colleagues , who used the process to demonstrate that dna was the genetic material of bacteria.in transformation, a cell takes up extraneous dna found in the environment and incorporates it into its genome (genetic material) through recombination. （一）mechanisms of transformat

11、iondonor dna binds to recipient cell at receptor site.one donor strands is degraded. donor strand is integrated into bacterial chromosome.after cell replication ,one cell is identical to original recipient the other carries the mutant gene.(二) transformation mappingcomplete mediumreplica platingtyra

12、+ cysc+tyra cysc+tyra+ cysctyra cyscminimalmediumminimal medium tyrosineminimal medium cysteine座位转 化 子 类 型trp2his2tyr1数目亲本类型重组类型重组值trp2-tyr1trp2-his2his2-tyr111940 3660 685 418 2600 107 11801194011801312011940107120471194036601560026001073660418678526001180366068581254181180107685239034. 01990567854

13、0. 020172812513. 0179902390trp2 his2 try1 trp2 his2 try1的转化子类型及重组值计算3413trp2his2tyr1the three possibility to engender prototrophmutation ?transformation ?nutrition reciprocate ?what were these colonies and how they did they arise ?二、conjugation(结合)conjugation: donor cells carrying specialized plasmi

14、dsestablish contact with and transfer dna to recipients.the experiment of lederberg and tatum bernard davis devised the u-shaped tube to answer these questions.the porous filter allowing the passageof liquid medium and dna but not bacteria cells mixed strain a with strain b in a u-shaped tube.no gro

15、wthno growththe experiment showed that direct cell-to-cell contact was necessary for the genetic exchange.conjugationn 试验材料：a品系：a-b+ t1s(met-bio-thr+leu+t1s)；b品系：a+b- t1r(met+bio+thr-leu-t1r)。n 试验方法：将a、b品系混合接种在基本培养基表面；短时间后喷t1杀死a品系，使其不能持续产生thr与leu供b品系持续生长。n 结果与结论：仍然出现原养型菌落。从而表明互养并非原养型菌落出现的原因，而可能发生了遗传

16、重组。营养互补？ in bacteria, conjugation is a one-way transfer with one strain acting as a donor and the other as a recipient. this was demonstrated by experiments in which one or the other strain was killed before mixing.hayes experiment:a: met thr leu thib: met thr leu thidonorreceptorkilledoccurredkille

17、dfaileda strainb strain(一) the f factor and conjugationf felectron micrograph of conjugation between an f (upper right) and f (lower left) cell with the f-pilus between them.(二) the conformation of the hfr cell whose chromosome was transferredcells whose chromosomes carry an integrated plasmid are c

18、alled hfr bacteria, because they produce a high frequency of recombinants for chromosomal genes in mating experiments with f strains.for a short while the f cell that has conjugated with an hfr cell has two copies, one copy of its own and one transferred in, the cell is a partial diploid, or a meroz

19、ygote.the new foreign dna is called exogenote; the host chromosome is called endogenote.(三) mapping genes by interrupted mating experimentshfr: thr leu azir tonr lac gal strsf: thr leu azis tons lac gal strrgenetic marker minutes after mating frequencythrleuazirtonrlacgal88.5911182510010090704025hfr

20、: thr leu azir tonr lac gal strs f: thr leu azis tons lac galstrrresults of the wollman-jocab interrupted mating experiment88.59111825thrleuazitonlacgalomap based on mating resultscomparisons of interrupted mating studies using different hfr strains confirm that the bacterial chrosomes is a circle (

21、四) recombination analysis of hfr crosses improve mapping accuracyhfr h strs lac ade f strr lac ade 根据中断杂交实验，知道lac 和 ade 是紧密连锁的，而且ade 是 lac之后进入f受体的。混合培养60minminimalstrf strr adelac也已进入flacadeadelacendogenoteexogenote把得到的重组子菌落影印培养在emb培养基上，鉴定能否利用乳糖。lac菌落为紫红色，lac菌落为粉红色。lacadestrsstrradelachfr染色体f染色体f染色体

22、strradelaclacadestrsstrradelacf染色体hfr染色体f染色体strradelac重组频率(rf)=%22%100adeadelacadelacadelacadelac用rf所测得的基因间距离与用中断杂交试验测定的t的基因间的距离基本上是一致的。两个值的比是rft201，即1 min等于20个图距单位(m.u.)。大肠杆菌染色体全长约90min，含3.6106核苷酸对，所以相当于1800 m.u.，1个m.u.约等于1600核苷酸对。三、 f-factor and sexduction (f-duction)the f-factor plus some bacteri

23、al dna is known asan f-factor sexduction 利用f因子将供体细胞的基因导入受体形成部分二倍体（partial diploid，merozygote）的过程。f因子转移细菌基因不同于hfr菌株，如： hfr: thr leu strs f: thr leu strr结果：筛选出f thr leu str r重组子 f thr leu strs f: thr leu strr结果：筛选出fthr leu str r重组子性导形成的部分二倍体为：lacflacfcomparing the conjugation cycles of f, hfr, fcells动

24、画演示during infection, a virus particle may incorporate a piece of the bacterial chromosome and introduce this piece of bacterial dna into other host cells during subsequent rounds of infection. the process by which viral particles transfer bacterial dna from one host cell to another is known as trans

25、duction. 四、转导( transduction )transduction: the bacterial gene transfer via bacteriophages1951年，j.lederberg 和 n.zinderphe trp tyrhis met与 phe trp tyrhismet原养型菌落10-5回复突变率为10-8 davis u型管实验证实是一种可滤性因子(fa)沙门氏属 salmonella志贺氏菌属 shigella变形杆菌属 proteus假单孢杆菌属 pseudomonas葡萄球菌属 staphylococcus芽孢杆菌属 bacillus转导可发生在转

26、导可发生在growhgrowh 具可滤性因子( filterable agent )存在minimal mediumfa与沙门氏菌的温和噬菌体p22在生物学特性上的一致性：(1)p22的侵染力与和fa的遗传力均可被脱氧核糖 核酸酶和胰蛋白酶破坏(2)用抗p22或用加热的处理均可使p22的侵染力和 fa的遗传能力失活，失活速率相同。(3)用抗p22的菌株作亲本进行上述混合培养，fa丧失侵染能力，不能出现重组体。(4)p22和fa在大小和质量上相同。1965年，k.ikeda 和j.tomizawa对e.coli的温和噬菌体的研究阐明了转导噬菌体是如何形成的。generalized transductionthe transduction process, which can result in the transfer of any bacterial gene between related strains of bacteria, is known as generalized transduction.transducing particle: a defective phage, one that carries bacterial dna rather than