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In the chapter of linkage, we have stated that the geneslocated in the same chromosome show linkage. Theselinked genes may either remain together during the process

of inheritance and, thus, showing complete linkage or they maybe segregated or separated during gametogenesis and, thus,displaying the incomplete linkage. The incomplete linkagetakes place due to the occurrence of new combinations orrecombinations of linked genes. The recombination in its turn isaccomplished through a process known as crossing over inwhich the non-sister chromatids of homologous chromosomesexchange the chromosomal parts or segments. In another words“the crossing over is a process that produces new combinations(recombinations) of genes by interchanging of correspondingsegments between non-sister chromatids of homologous chro-mosomes”. The chromatins resulting from such interchanges ofchromosomal parts are known as cross overs. The term cross-ing over was coined by Morgan. No crossing over occurs inmale Drosophila and female silk worm, Bombyx mori. Certainexternal agents such as heat shock, chemicals, radiations, etc.,have profound effect on crossing over.

Characteristics of Crossing Over1. Crossing over or recombination occurs at two levels

(i) at gross chromosomal level, called chromosomal crossingover and (ii) at DNA level, called genetic recombination.

2. A reciprocal exchange of material between homolo-gous chromosomes in heterozygotes is reflected in crossingover.

Crossing Over

C H A P T E R

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Prophase – I stage showing crossingover.

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Fig. 8.1. Somatic or mitotic crossing over in Drosophila. A—Thorax of a fruit fly heterozygous for y (yellowbody) and sn (singed hairs and bristles) with a twin spot; B—Mechanism of crossing over leading tohomozygosity for distal genes.

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3. The crossing over results basically from an exchange of genetic material between non-sisterchromatids by break-and-exchange following replication.

4. The frequency of crossing over appears to be closely related to physical distance betweengenes on chromosome and serves as a tool in constructing genetic maps of chromosomes.

TYPES OF CROSSING OVERAccording to its occurrence in the somatic or germ cells following two types of crossing over

have been recognized :

1. Somatic or Mitotic Crossing Over

When the process of crossing over occursin the chromosomes of body or somatic cells ofan organism during the mitotic cell division it isknown as somatic or mitotic crossing over. Thesomatic crossing over is rare in its occurrenceand it has no genetical significance. The somaticor mitotic crossing over has been reported in thebody or somatic cells of Drosophila by CurtStern and in the fungus Aspergillus nidulans byG. Pontecorvo.

Example. Stern (1936) observed thatDrosophila females which are heterozygous forsex-linked recessive genes, yellow body colour(Y) and singed (=burned or scorched) hairs andbristles (sn), are phenotypically like wild typesbut occasionally show yellow spots on theirbody. Homozygous recessive individuals for these genes are yellow bodied and with bent bluntedbristles. Normally, all the cells of a heterozygous should be normal and should not express either of

Demonstration of sister chromatid exchanges inmitotic chromosomes.

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CROSSING OVER 95

these genes. Yellow skin spots can appear only if cells are homozygous for y, indicating thereby thatwhenever yellow spots appear in heterozygous flies, there is prior crossing over between Y and snalleles. Stern found that equal sections of singed and yellow tissues lie adjacent to each other or formtwin spots. This indicate that such tissues arise as a result of mitotic crossing over, rather than due tomutation. Somatic or mitotic crossing over occurs at a four strand stage and during this process thereis pairing of homologous chromosomes (Fig. 8.1).

2. Germinal or Meiotic Crossing OverUsually the crossing over occurs in germinal cells during the gametogenesis in which the meiotic

cell division takes place. This type of crossing over is known as germinal or meiotic crossing over.The meiotic crossing over is universal in its occurrence and is of great genetic significance.

MECHANISM OF MEIOTIC CROSSING OVERThe process of crossing over includes following stages in it, viz., synapsis, duplication of

chromosomes, crossing over and terminalization. The chromosomes which tend to undergo recombi-nation due to meiotic crossing over necessarily complete two functions : 1. 99.7 per cent replicationof DNA and 75 per cent synthesis of histones, both of which take place prior to onset of prophase 1,and 2. attachment of each chromosome by its both ends (telomeres) to the nuclear envelope (i.e., tonuclear lamina) via the specialized structure, called attachment plaques (see Suzuki et al., 1986).This event occurs during the leptotene stage of prophase I and though each chromosome at this stageis visually long and thin thread, but contains material of two sister chromatids (i.e., two DNA moleculesplus almost duplicated amount of histones).

1. SynapsisSynapsis or intimate pairing between

the two homologous chromosomes (one ma-ternal and another paternal) is initiated duringzygotene stage of prophase I of meiosis I.Synapsis often starts when the homologousends of the two chromosomes are broughttogether on the nuclear envelope and it con-tinues inward in a zipper-like manner fromboth ends, aligning the two homologous chro-mosomes side by side (e.g., mammals). Inother cases, synapsis may begin in internalregions of the chromosomes and proceedtoward the ends, producing the same type of alignment. By synapsis each gene is, thus, brought intojuxtaposition (=being side by side) with its homologous gene on the opposite chromosome. Thus,synapsis is the phase of prolonged and close contact of homologous chromosomes due to attractionbetween two exactly identical or homologous regions or chromomeres. The resultant pairs ofhomologous chromosomes are called bivalents. The phenomenon of synapsis has always intriguedcytogeneticists. While Darlington tried to explain the cause of synapsis by proposing his precocitytheory, Moses identified a factor in the formation of synaptonemal complex which aids in synapsis.

(a) Precocity theory. To explain the question that why do homologous chromosomes, duringsynapsis, approach each other from a considerable distance and become closely associated, a Britishcytologist C.D. Darlington in 1937, proposed the precocity theory of meiosis which embraces wellthe cause of synapsis in it. According to the precocity theory of meiosis, chromosomes enter intomeiotic prophase I, in contrast to mitotic prophase, as unreplicated structures, each of which consistingof a single chromatid which he considered as unbalanced or unsaturated state in electrostatical

Telomeres (shown here in yellow colour).

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relations. In order to become saturated or balanced the chromosomes must pair. Thus, the sequencesof meiotic pairing or synapsis are determined precociously.

Precocity theory is now untenable since it erroneously stressed upon the idea that DNA synthesisor chromosome duplication takes place later in pachytene or diplotene and then results in separationof homologous chromosomes.

(b) Synaptonemal complex. Montrose J. Moses (1956) has revealed a highly organizedstructure of filaments called syn-aptonemal complex in betweenthe paired chromosomes of zygo-tene and pachytene stages in cray-fish by electron microscopy. Syn-aptonemal complex has also beenobserved in a wide variety of spe-cies of plants and animals.

In electron-micrographs thesynaptonemal complex appears asthree parallel dense lines that lieequally spaced in a plane and areflanked by chromatin. The ele-ments of two lateral lines usuallyappear densest, while the elementof central line is of variable promi-nence. Some fine transversestrands also cross between lateralelements, connecting them withthe central element. Though, the morphology of lateral and central elements may vary from species tospecies, but the basic structure and the spacing of the synaptonemal complex is constant within thespecies.

Cytochemical studies have shown that the lateral elements of synaptonemal complex are rich inDNA, RNA and proteins, but that the central element of it contains mainly RNA, protein and little DNA.

Functions of synaptonemal complex. The synaptonemal complex is found to be concomitantof both chiasma formation and crossing over. For instance, the synaptonemal complex may servecrossing over by facilitating effective synapsis in one or more of the following ways—(a) to maintainpairing in fixed state for an extended period, (b) to provide a structural framework within whichmolecular recombination may occur, and (c) to segregate recombination DNA from the bulk of thechromosomal DNA.

Robert King (1970) suggested that the synaptonemal complex may orient the non-sisterchromatids of homologs in a manner to facilitate enzymatically induced exchanges between their DNAmolecules. More recently, D.A. Comings and T.A. Okada (1971), have shown electron microscopi-cally that synapsis occurs at two levels one at chromosomal level and the other at the molecular level.According to them, the synaptonemal complex pulls homologous chromosomes into approximateassociation with each other but plays no role in molecular pairing of DNA strand.

Recently, each event of recombination due to crossing over is found to be performed by a largeprotein assembly of 90 nm diameter, called recombination nodule, which is placed on thesynaptonemal complex (Carpenter, 1977, 1987). Each nodule marks the site of a large multienzyme‘recombination machine’ which brings local regions of DNA on maternal and paternal chromatidstogether across the 100-nm wide synaptonemal complex.

Fig. 8.2. A typical synaptonemal complex (after Alberts et al., 1989).

recombination nodule lateral elements

chromatin of sisterchromatids 1 and 2

(paternal)

centralelements chromatin of sister chromatids

3 and 4 (maternal)

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2. Duplication of ChromosomesThe synapsis is followed by duplication of chromosomes (in pachytene). During this stage, each

homologous chromosomeof bivalents splits longitu-dinally and form two identi-cal sister chromatids whichremain held together by anunsplitted centromere. Thelongitudinal splitting ofchromosomes is achieved bythe separation of already du-plicated DNA moleculesalong with certain chromo-somal proteins. At this stage each bivalent contains four chromatids, so it is known as tetrad.

3. Crossing Over by Breakage and UnionIt is well evident that crossing over occurs in the homologous chromosomes only during the four

stranded or tetrad stage. Homologues continue to stay in synapsis for days during pachytene stage andchromosomal crossing over occurs due to exchange of chromosomal material between non-sisterchromatids of each tetrad. In pachytene, the recombination nodules become visible between synapsedchromosomes.

During the process of crossing over, two non-sister chromatids first break at the correspondingpoints due to the activity of a nuclear enzyme known as endonuclease (Stern and Hotta, 1969). Thena segment on one side of each break connects with a segment on the opposite side of the break, so thatthe two non-sister chromatids cross each other. At this stage .3 per cent synthesis of DNA (i.e., P–DNAreplication) occurs to fill the gap. The fusion of chromosomal segments with that of opposite one takesplace due to the action of an enzyme known as ligase (Stern and Hotta, 1969). The crossing of twochromatids is known as chiasma (Gr., chiasma=cross) formation. The crossing over, thus, includesthe breaking of chromatid segments, their transposition and fusion.

Chiasma frequency or percentage of crossing over. The crossing over may take place atseveral points in one tetrad and may result in the formation of several chiasmata. The number ofchiasmata depends on the length of the chromosomes because the longer the chromosome the greaterthe number of chiasmata. In a species each chromosome has a characteristic number of chiasmata. Thefrequency by which a chiasmata occurs between any two genetic loci has also a characteristicprobability. The more apart two genes are located on a chromosome, the greater the opportunity fora chiasma to occur between them. The closer two genes are linked lesser the chances for a chiasmaoccurring between them.

4. TerminalisationAfter the occurrence of process of crossing over, the non-sister chromatids start to repel each

other because the force of synapsis attraction between them decreases. During diplotene, desynapsisbegins, synaptonemal complex dissolves and two homologous chromosomes in a bivalent are pulledaway from each other. During diakinesis, chromosomes detaches from the nuclear envelope and eachbivalent is clearly seen to contain four separate chromatids with each pair of sister chromatids linkedat their centromeres, while non-sister chromatids that have crossed over are linked by chiasmata. Thechromatids separate progressively from the centromere towards the chiasma and the chiasma itselfmoves in a zipper fashion towards the end of the tetrad (Fig. 8.4). The movement of chiasma is knownas terminalisation. Due to the terminalisation the homologous chromosomes are separated com-pletely.

Fig. 8.3. Diagram showing the mechanism of crossing over.

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KINDS OF CROSSING OVERAccording to the number of chiasma following types of

crossing over have been described.1. Single crossing over. When the chiasma occurs only at

one point of the chromosome pair then the crossing over is knownas single crossing over. The single crossing over produces twocross over chromatids and two non-cross over chromatids.

2. Double crossing over. When the chiasmata occur at twopoints in the same chromosome, the phenomenon is known asdouble crossing over. In the double crossing over, the formationof each chiasma is independent of the other and in it four possibleclasses of recombination occur. In the double crossing overfollowing two types of chiasma may be formed :

(i) Reciprocal chiasma. In the reciprocal chiasma the sametwo chromatids are involved in the second chiasma as in the first.Thus, the second chiasma restores the order which was changed bythe first chiasma and it produces two non-cross over chromatids.The reciprocal chiasma occurs in two strand double crossing overin which out of four chromatids only two are involved in thedouble crossing over.

(ii) Complimentary chiasma. When both the chromatidstaking part in the second chiasma are different from those chroma-tids involved in the first chiasma, the chiasma is known ascomplimentary chiasma. The complimentary chiasma producesfour single cross overs but no non-cross over. The complimentarychiasma occurs when three or four chromatids of the tetradundergo the crossing over.

3. Multiple crossing over. When crossing over takes placeat more than two places in the same chromosome pair then suchcrossing over is known as multiple crossing over. The multiple crossing over occurs rarely.

THEORIES ABOUT THE MECHANISM OF CROSSING OVERFollowing theories have been propounded to explain the mechanism of crossing over :1. Duplication theory. This theory was proposed by John Belling (1928) while studying

meiosis in some plant species. Belling believed that crossing over might occur during duplication ofhomologous chromosomes and might broughtabout due to novel attachments formed betweennewly synthesized genes. He visualized genes asbeads (described as chromomeres), connectedby non-genic linking elements—theinterchromomeric regions. During duplicationof chromosomes, initially the chromomeres areduplicated and the newly formed chromomeresremain tightly juxtaposed to the old ones. Wheninterchromomeric regions are synthesized to jointhese new genes or chromomeres, they may switchfrom a newly synthesized chromomere on one

Fig. 8.4. Diagram showing theterminalisation.

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The photograph illustrates several chiasmata found ina tetrad isolated during first meiotic prophase stage.

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