Lecture 1 Genetics PDF

Genetics Lecture 1 Dr/ zayed Prepared & presented by Dr/ Basant Yosri Lec 1 genetics Dr/ Zayed Dr/ Basant Yosri Chromosomes and Cell Division INTRODUCTION DNA can be...

Genetics Lecture 1 Dr/ zayed Prepared & presented by Dr/ Basant Yosri Lec 1 genetics Dr/ Zayed Dr/ Basant Yosri Chromosomes and Cell Division INTRODUCTION DNA can be regarded as the basic template that provides a blueprint for the formation and maintenance of an organism, inform of chromosomes chromosomes made up of tightly coiled long chains of genes which can be visualized during cell division using a light microscope, which appear as threadlike structures or ‘colored bodies’. The word chromosome is derived from the Greek chroma (= color) and soma (= body). enable the transmission of genetic information from one generation to the next. Their behavior at somatic cell division in mitosis provides a means of ensuring that each daughter cell retains its own complete genetic complement. Chromosomes are quite literally the vehicles that facilitate reproduction and the maintenance of a species. As their behavior during gamete formation in meiosis enables each mature ovum and sperm to contain a unique single set of parental genes cytogenetics→The study of chromosomes and cell division Human chromosomes At the submicroscopic level chromosomes consist of an extremely elaborate complex, made up of supercoils of DNA, which has been likened to the tightly coiled network of wiring seen in a solenoid Note→ most of chromosome structure has been gained using light microscope how? Because of using special stain take up by DNA enable each chromosome to be identified ( best time doing that is during cell division → chromosome is maximally contracted and the constituent genes can no longer be transcribed. each chromosome can be seen to consist of two identical strands known as chromatids, or sister chromatids, which are the result of DNA replication having taken place during the S (synthesis) phase of the cell cycle centromere it is the sister chromatids which can be seen to be joined at a primary constriction consist of several hundred kilobases of repetitive DNA and are responsible for the movement of chromosomes at cell division Each centromere divides the chromosome into short p (= petite) and long arms q (‘g’ = grande), Telomere The tip of each chromosome arm Telomeres play a crucial role in sealing the ends of chromosomes and maintaining their structural integrity. Telomeres have been highly conserved throughout evolution and in humans they consist of many tandem repeats of a TTAGGG sequence. During DNA replication, an enzyme known as telomerase replaces the 5′ end of the long strand, which would otherwise become progressively shorter until a critical length was reached when the cell could no longer divide and thus became senescent. in some tumors, increased telomerase activity has been implicated as a cause of abnormally prolonged cell survival. But most cells being unable to undergo more than 50 to 60 divisions. Lec 1 genetics Dr/ Zayed Dr/ Basant Yosri Under the electron microscope chromosomes can be seen to have a rounded and rather irregular morphology Electron micrograph of human chromosomes showing the centromeres and well-defined chromatids. Morphologically chromosomes are classified according to: 1. the position of the centromere located centrally→the chromosome is metacentric Located terminally →the chromosome is acrocentric Located intermediate→the chromosome is submetacentric. Acro-centric chromosomes sometimes have stalk-like appendages called satellites that form the nucleolus of the resting interphase cell and contain multiple repeat copies of the genes for ribosomal RNA. 2. overall length 3. presence or absence of satellites note: early pioneers of cytogenetics were able to identify most individual chromosomes, or at least subdivide them into groups labeled A to G on the basis of overall morphology (A, 1–3; B, 4–5; C, 6–12 X; D, 13–15; E, 16–18; F, 19–20; G, 21–22 1 Y). humans the normal cell nucleus contains 46 chromosomes, made up of 22 pairs of autosomes and a single pair of sex chromosomes—XX in the female and XY in the male. One member of each of these pairs is derived from each parent Somatic cells may have: 1. diploid complement of 46 chromosomes 2. gametes (ova and sperm) have a haploid complement of 23 chromosomes. 3. Members of a pair of chromosomes are known as homologs. Chromatin combination of DNA and histone proteins that comprise chromosomes, exists in two main forms. 1. Euchromatin stains lightly and consists of genes that are actively expressed. 2. heterochromatin stains darkly and is made up largely of inactive, unexpressed, repetitive DNA. Lec 1 genetics Dr/ Zayed Dr/ Basant Yosri The Sex Chromosomes (X and Y chromosomes) It is called sex chromosome because of their crucial role in sex determination in some insects this chromosome is present in some gametes but not in others. In these insects the male has only one sex chromosome (X), whereas the female has two (XX) In humans, and in most mammals, both the male and the female have two sex chromosomes—XX in the female and XY in the male. In the female each ovum carries an X chromosome, whereas in the male each sperm carries either an X or a Y chromosome. there is a roughly equal chance of either an X-bearing sperm or a Y-bearing sperm fertilizing an ovum, the numbers of male and female conceptions are approximately equal (Figure 3.3). In fact, slightly more male babies are born than females, although during childhood and adult life the sex ratio evens out at 1 : 1. The Y chromosome is much smaller than the X and carries only a few genes of functional importance, most notably the testis-determining factor, known as SRY. Other genes on the Y chromosome are known to be important in maintaining spermatogenesis. Punnett square showing sex chromosome combinations for male and female gametes. Lec 1 genetics Dr/ Zayed Dr/ Basant Yosri Methods of Chromosome Analysis karyotyping universally employed in cytogenetic laboratories to analyze the chromosome constitution of an individual term used to describe a photomicrograph of an individual’s chromosomes, arranged in a standard manner. Steps of karyotyping 1. Chromosome Preparation Source of sample→ Any tissue with living nucleated cells that undergo division a) Most commonly circulating lymphocytes from peripheral blood b) Skin c) bone marrow d) chorionic villi (7) e) cells from amniotic fluid (amniocytes). stained with a DNA binding dye—also known as ‘Giemsa’—that gives each chromosome a characteristic and reproducible pattern of light and dark bands (1) sample is added to a (6) small volume of chromosomes nutrient medium are treated with containing trypsin, which phytohemagglutinin, denatures their which stimulates T- protein content. lymphocytes to divide. (5) then fixed, mounted on a (2) slide and stained ready for analysis Cells are cultured under sterile conditions at 37°C for about 3 days (4) Hypotonic saline is then added, which (3) causes the blood colchicine is then added to each culture. This drug has the cells to lyse and extremely useful property of preventing formation of the spindle, results in thereby arresting cell division during metaphase, the time when the spreading of the chromosomes are maximally condensed and therefore most visible. chromosomes Lec 1 genetics Dr/ Zayed Dr/ Basant Yosri 2. Chromosome Banding G banding generally provides high-quality chromosome analysis with approximately 400 to 500 bands per haploid set. Each of these bands corresponds on average to approximately 6000 to 8000 kilobases (kb) (i.e., 6 to 8 megabases [mb]) of DNA. High-resolution banding of the chromosomes at an earlier stage of mitosis, such as prophase or prometaphase, provides greater sensitivity with up to 800 bands per haploid set, but is much more demanding technically inhibiting cell division with an agent such as methotrexate or thymidine. Folic acid or deoxycytidine is added to the culture medium, releasing the cells into mitosis. Colchicine is then added at a specific time interval, when a higher proportion of cells will be in prometaphase and the chromosomes will not be fully contracted, giving a more detailed banding pattern. 3.Karyotype Analysis a) metaphase spreads→ counting the number of chromosomes present in a specified number of cells b) idiogram→ banding pattern of each chromosome is specific and can be shown in the form of a stylized ideal karyotype The cytogeneticist analyzes each pair of homologous chromosomes, either directly by looking down the microscope or using an image capture system to photograph the chromosomes and arrange them in the form of a karyogram A G-banded metaphase spread. Lec 1 genetics Dr/ Zayed Dr/ Basant Yosri Molecular Cytogenetics (Fluorescent In-Situ Hybridization) This diagnostic tool combines conventional cytogenetics with molecular genetic technology. It is unique ability of a portion of single-stranded DNA (i.e., a probe) to anneal with its complementary target sequence on a metaphase chromosome, interphase nucleus or extended chromatin fiber. In fluorescent in-situ hybridization (FISH) DNA probe is labeled with a fluorochrome which, after hybridization with the patient’s sample, allows the region where hybridization has occurred to be visualized using a fluorescence microscope. Different Types of FISH Probe Centromeric These consist of repetitive DNA sequences found in and Probes around the centromere of a specific chromosome They were the original probes used for rapid interphase FISH diagnosis of the common aneuploidy syndromes from a prenatal diagnostic sample of chorionic villi until it was superseded by quantitative fluorescent polymerase chain reaction. Chromosome- These are specific for a particular single locus which Specific Unique- can be used to identify submicroscopic deletions and Sequence Probes duplications causing microdeletion syndromes Another application is the use of an interphase FISH probe to identify HER2 overexpression in breast tumors to identify patients likely to benefit from Herceptin treatment. Metaphase image of Williams (ELN) region probe (Vysis), chromosome band 7q11.23, showing the deletion associated with Williams syndrome. The normal chromosome has signals for the control probe (green) and the ELN gene probe (orange), but the deleted chromosome shows only the control probe signal. Whole- These consist of a cocktail of probes obtained from Chromosome Paint different parts of a particular chromosome. Probes When this mixture of probes is used together in a single hybridization, the entire relevant chromosome fluoresces (i.e., is ‘painted’ Chromosome painting is useful for characterizing complex rearrangements, such as subtle translocations and for identifying the origin of additional chromosome material, such as small supernumerary markers or rings. Chromosome painting showing a reciprocal translocation involving chromosomes 3 (red) and 20 (green). Lec 1 genetics Dr/ Zayed Dr/ Basant Yosri Chromosome Nomenclature By convention each chromosome arm is divided into regions and each region is subdivided into bands, numbering always from the centromere outwards A given point on a chromosome is designated by the chromosome number, the arm (p or q), the region, and the band (e.g., 15q12). Sometimes the word region is omitted, so that 15q12 would be referred to simply as band 12 on the long arm of chromosome 15. A shorthand notation system exists for the description of chromosome abnormalities Normal male and female karyotypes are depicted as 46,XY and 46,XX, respectively. A male with Down syndrome as a result of trisomy 21 would be represented as 47,XY,+21, whereas a female with a deletion of the short arm of one number 5 chromosome (cri du chat syndrome) would be represented as 46,XX,del(5p). A chromosome report reading 46,XY,t(2;4) (p23;q25) would indicate a male with a reciprocal translocation involving the short arm of chromosome 2 at region 2 band 3 and the long arm of chromosome 4 at X chromosome showing the region 2 band 5. short and long arms each subdivided into regions and bands. Lec 1 genetics Dr/ Zayed Dr/ Basant Yosri Cell Division Mitosis This process of somatic cell division, during which the nucleus also divides, is known as mitosis. During mitosis each chromosome divides into two daughter chromosomes, one of which segregates into each daughter cell. Consequently, the number of chromosomes per nucleus remains unchanged. Mitosis is the process whereby each of pairs of chromatids(chromosome consists of two identical sister chromatids as a result of DNA replication having taken place during the S phase of the cell cycle )separates and disperses into separate daughter cells. Mitosis is a continuous process that usually lasts 1 to 2 hours Divided into five distinct stages. I. Prophase During the initial stage of prophase, the chromosomes condense and the mitotic spindle begins to form. Two centrioles form in each cell, from which microtubules radiate as the centrioles move toward opposite poles of the cell. II. Prometaphase During prometaphase the nuclear membrane begins to disintegrate, allowing the chromosomes to spread around the cell. Each chromosome becomes attached at its centromere to a microtubule of the mitotic spindle. III. Metaphase In metaphase the chromosomes become aligned along the equatorial plane or plate of the cell, where each chromosome is attached to the centriole by a microtubule forming the mature spindle. At this point the chromosomes are maximally contracted and, therefore, most easily visible. Each chromosome resembles the letter X in shape, as the chromatids of each chromosome have separated longitudinally but remain attached at the centromere, which has not yet undergone division. IV. Anaphase In anaphase the centromere of each chromosome divides longitudinally and the two daughter chromatids separate to opposite poles of the cell. V. Telophase By telophase the chromatids, which are now independent chromosomes consisting of a single double helix, have separated completely and the two groups of daughter chromosomes each become enveloped in a new nuclear membrane. The cell cytoplasm also separates (cytokinesis), resulting in the formation of two new daughter cells, each of which contains a complete diploid chromosome complement. Lec 1 genetics Dr/ Zayed Dr/ Basant Yosri The Cell Cycle Interphase of the cell cycle→ The period between successive mitoses rapidly dividing cells the interphase: a. lasts for between 16 and 24 hours b. commences with the G1 (G = gap) phase during which the chromosomes become thin and extended. c. Very variable in length and is responsible for the variation in generation time between different cell populations. stopped dividing, such as neurons the interphase: a. cell said to entered a noncyclic stage known as G0 b. The G1 phase is followed by the S phase (S = synthesis) c. DNA replication occurs and the chromatin of each chromosome is Inactive X Chromosome replicated. It commences at multiple it is one of the X chromosomes is always late points on a chromosome in replicating ,when Homologous pairs of d. results in the formation of two chromosomes usually replicate is synchrony. chromatids, giving each chromosome It forms the sex chromatin or so-called Barr its characteristic X-shaped body, which can be visualized during configuration. interphase in female somatic cells. This used to be the basis of a rather Note: Interphase is completed by a relatively short G2 phase during which the unsatisfactory means of sex determination chromosomes begin to condense in based on analysis of cells obtained by scraping preparation for the next mitotic division. the buccal mucosa—a ‘buccal smear’. Lec 1 genetics Dr/ Zayed Dr/ Basant Yosri Meiosis called reduction division Because it is during the first meiotic division that the chromosome number is halved. Meiosis Mitosis the process of nuclear division that occurs during process of somatic cell division, during which the the final stage of gamete formation. nucleus also divides the diploid count is halved so that each mature results in each daughter cell having a diploid gamete receives a haploid complement of 23 chromosome complement (46). chromosomes. occurs only at the final division of gamete takes place in somatic cells and during the early maturation. cell divisions in gamete formation. can be considered as two cell divisions known as occurs as a one-step process meiosis I and meiosis II, each of which can be considered as having prophase, metaphase, anaphase, and telophase stages, as in mitosis Meiosis 1 phases 1.Prophase I Chromosomes enter this stage already split longitudinally into two chromatids joined at the centromere. Homologous chromosomes pair and, with the exception of the X and Y chromosomes in male meiosis, exchange of homologous segments occurs between non-sister chromatids; that is, chromatids from each of the pair of homologous chromosomes. This exchange of homologous segments between chromatids occurs as a result of a process known as crossing over or recombination. The importance of crossing over in linkage analysis and risk calculation is considered later During prophase I in the male, pairing occurs between homologous segments of the X and Y chromosomes at the tip of their short arms, with this portion of each chromosome being known as the pseudo autosomal region. The prophase stage of meiosis I is relatively lengthy and can be subdivided into five stages. Leptotene. The chromosomes become visible as they start to condense. Zygotene. Homologous chromosomes align directly opposite each other, a process known as synapsis, and are held together at several points along their length by filamentous structures known as synaptonemal complexes Pachytene. Each pair of homologous chromosomes, known as a bivalent, becomes tightly coiled. Crossing over occurs, during which homologous regions of DNA are exchanged between chromatids. Diplotene. The homologous recombinant chromosomes now begin to separate but remain attached at the points where crossing over has occurred. These are known as chiasmata. On average, small, medium, and large chromosomes have one, two, and three chiasmata, respectively, giving an overall total of approximately 40 recombination events per meiosis per gamete. Diakinesis. Separation of the homologous chromosome pairs proceeds as the chromosomes become maximally condensed.v Lec 1 genetics Dr/ Zayed Dr/ Basant Yosri 2.Metaphase I The nuclear membrane disappears and the chromosomes become aligned on the equatorial plane of the cell where they have become attached to the spindle, as in metaphase of mitosis. 3.Anaphase I The chromosomes now separate to opposite poles of the cell as the spindle contracts. 4.Telophase I Each set of haploid chromosomes has now separated completely to opposite ends of the cell, which cleaves into two new daughter gametes, so-called secondary spermatocytes or oocytes. Meiosis II This is essentially the same as an ordinary mitotic division. Each chromosome, which exists as a pair of chromatids, becomes aligned along the equatorial plane and then splits longitudinally, leading to the formation of two new daughter gametes, known as spermatids or ova. The Consequences of Meiosis Regards reproduction and the maintenance of the species→ provide 2 objectives I. it facilitates halving of the diploid number of chromosomes so that each child receives half of its chromosome complement from each parent. II. it provides an extraordinary potential for generating genetic diversity. This is achieved in two ways: A. first When the bivalents separate during prophase of meiosis I, they do so independently of one another. This is consistent with Mendel’s third law. Consequently each gamete receives a selection of parental chromosomes. The likelihood that any two gametes from an individual will contain exactly the same chromosomes is 1 in 223, or approximately 1 in 8 million. B. second As a result of crossing over, each chromatid usually contains portions of DNA derived from both parental homologous chromosomes. A large chromosome typically consists of three or more segments of alternating parental origin. The ensuing probability that any two gametes will have an identical genome is therefore infinitesimally small. This dispersion of DNA into different gametes is sometimes referred to as gene shuffling.  Lec 1 genetics Dr/ Zayed Dr/ Basant Yosri Gametogenesis The process of gametogenesis shows fundamental differences in males and females, These have quite distinct clinical consequences if errors occur. Mature ova develop from oogonia by a complex spermatogenesis is a relatively rapid series of intermediate steps. process with an average duration of 60 Oogonia themselves originate from primordial germ to 65 days. cells by a process involving 20 to 30 mitotic divisions At puberty spermatogonia, which will that occur during the first few months of embryonic already have undergone approximately life. 30 mitotic divisions, begin to mature into By the completion of embryogenesis at 3 months of primary spermatocytes which enter intrauterine life, the oogonia have begun to mature meiosis I and emerge as haploid into primary oocytes that start to undergo meiosis. secondary spermatocytes. At birth all of the primary oocytes have entered a These then undergo the second meiotic phase of maturation arrest, known as dictyotene, in division to form spermatids, which in which they remain suspended until meiosis I is turn develop without any subsequent cell completed at the time of ovulation, when a single division into mature spermatozoa, of secondary oocyte is formed. which 100 to 200 million are present in This receives most of the cytoplasm. each ejaculate. The other daughter cell from the first meiotic Spermatogenesis is a continuous division consists largely of a nucleus and is known as process involving many mitotic divisions, a polar body. possibly as many as 20 to 25 per annum, Meiosis II then commences, during which so that mature spermatozoa produced fertilization can occur. This second meiotic division by a man of 50 years or older could well results in the formation of a further polar body have undergone several hundred mitotic divisions.  Lec 1 genetics Dr/ Zayed Dr/ Basant Yosri the very lengthy interval between the onset of meiosis and its eventual completion, up to 50 years later, accounts for the well documented increased The observed paternal age effect for incidence of chromosome abnormalities in the new dominant mutations (p. 69) is offspring of older mothers consistent with the concept that many mutations arise as a consequence of The accumulating effects of ‘wear and tear’ on the DNA copy errors occurring during primary oocyte during the dictyotene phase mitosis. probably damage the cell’s spindle formation and repair mechanisms, thereby predisposing to non- disjunction