Molecular Genetics

Molecular genetics is the study of molecules and mechanisms involved in genetic inheritance. Archival information molecules are long polymers of deoxyribonucleic acid (DNA) comprising bases in specific sequence. The bases adenine (A), thymine (T), cytosine (C), and guanine (G), function as codon triplets – sequences of three bases that code for specific amino acids or for translation initiation (start codons) or termination (stop codons). Uracil is substituted for thymine in RNA.

The segments of DNA that contain protein-coding instructions are called genes, and these gene sequences comprise a portion of the total genome of a cell. The genome includes both the genes (coding-sequences, domains) and the non-coding sequences – both exons, which include open reading frames, and introns.

Because the 64 possible combinations of GATC code for only the 20 amino acids commonly found in proteins, the code is 'degenerate' (redundant) with more than one triplet combination coding for each amino acid. (This code reduncancy provides hereditary stability by reducing mutation mistakes.) The double helix of DNA comprises paired nucleotide strands with bases hydrogen bonded to complementary bases in the adjacent chain. Adenine pairs with thymine or uracil (A-TU), and cytosine pairs with guanine (CG).

During cellular reproduction, strands of archival DNA are copied or replicated. Transcription is the first step in gene expression – DNA instructions are converted into mRNA codons, rRNAs, miRNAs, and tRNAs. Coding instructions of nucleotide sequences in archival DNA, which have been transcribed and processed into mRNAs are translated into polypeptides and proteins at cytoplasmic ribosomes. Translation is the ultimate step in gene expression, in which archival genetic instructions are converted into specified sequences of amino acids in peptides, polypeptides, and proteins.

In prokaryotic cells – without a nuclear membrane – translation into polypeptides and proteins may begin prior to termination of transcription. The molecular genetics of eukaryotic cells is more complicated than that of prokaryotes. Various molecules of ribonucleic acid (RNA) participate in the transcription of the DNA code into processed mRNA in a series of RNA processing stages including capping, polyadenylation, and pre-mRNA splicing.Following pre-mRNA processing, RNAs undergo extranuclear transfer. Mature RNAs may undergo post-transcriptional modulation (via miRNAs) before translation of the archival DNA instructions into specific sequences of amino acids in the polypeptides and proteins that participate in cellular function and structure. Transfer RNAs (tRNA) deliver specific amino acids to the cytoplasmic ribosomes along the rough endoplasmic reticulum. Ribosomal RNAs participate in assembly of polypeptides and proteins at ribosomes. Here RNAs serve as ribozymes – non-protein enzymes.

A number of processes are involved in control of cellular function through the maintenance of accuracy of genetic inheritance – damage to DNA is repaired, and faulty RNA is destroyed.

DNA damage may result from replication errors, incorporation of mismatched nucleotides (substitution errors – transitions and transversions), oxygen radicals, hydroxyl radicals, ionizing or ultraviolet radiation, toxins, alkylating agents, and chemotherapy agents. A number of vital mechanisms repair DNA damage to bases (including C to T, C to U, and T U mismatch) and to strands, including double strand breaks. All organisms, prokaryotic and eukaryotic, utilize at least three enzymatic excision-repair mechanisms for damaged bases: base excision repair, mismatch repair, and nucleotide excision repair.

Given the importance of mRNA as an information-carrying molecule, faulty pre-mRNAs and mRNAs must be eliminated – they are destroyed by nonsense-mediated decay or nonstop decay:
1. A pre-mRNA made from a mutant gene usually has an exon junction complex (EJC) in the wrong position. This error activates nonsense-mediated decay (NMD) and destroys the pre-mRNA before it can be used to make flawed proteins. There are at least two kinds of NMD: one requires the protein UPF2 and the other does not.
2. Nonstop decay is mRNA turnover mechanism that has none of the properties of normal mRNA turnover or of NMD. A multi-enzyme complex called the exosome is important for nonstop decay. The exosome is the site for binding of a specific adapter protein called Ski7p. Nonstop decay shares none of the enzymes required for nonsense-mediated decay.

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Important features of the genome and proteome

Important features of the genome and proteome:
1) Segments of nucleic acid that code for a useful sequence can be re-used, and need not arise de novo for each protein in an organism’s proteome.
2) The original genetic code probably employed 2 not 3 bases to code for each amino acid, and there were probably less than 20 constituent amino acids in primordial proteomes.
3) Many codons – the 2 or 3 base coding sequences – overlap with one another. That is, several sequences may code for a single amino acid within the polypeptide. This redundancy feature protects the proteome from random point mutations in the genome. For example, these are the triplet codons for arginine: CGU CGC CGA CGG AGA AGG
4) The function of proteins depends upon their tertiary and quaternary (3D) structure, and not on the primary (amino acid sequence) structure.
5) Homologous proteins, those that perform similar functions in different organisms, share sequences that are evolutionarily invariant, or conserved. Other amino acids may be substituted into the sequences whose sole function is to connect the important sequences without altering the 3D configuration or function of a protein.
6) Many small RNAs are important as enzymes and in epigenetic regulation – these RNAs are coded for by segments of DNA that do not code for proteins. Translation of RNA to protein is orchestrated by RNA and proteins.
7) Because the sequences of bases in RNA and one strand of DNA are complementary, the sequences of bases in RNA are equivalent to those in the other strand of DNA (with RNA's U switched to DNA's T). This means that RNA could have provided the template for encoding of its own sequences, thus eliminating any need for re-invention of a DNA code from which to transcribe RNAs.
8) The enormous number of different sequences of bases in the hypothetical 26 base RNA strand in the example demonstrates the possible ‘experiments’ that could be performed in a primordial soup mix. Remember that the example merely examined permutations and combinations for a 26 base nucleic acid polymer.

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Conserved & Consensus

In general, a consensus sequence is that idealized sequence in which each position represents the base/amino acid most often found when many sequences are compared. A genetic consensus sequence is a sequence of nucleotides that is common to different genes or genomes. There may be some variations but such sequences show considerable similarity. So, a consensus sequence is the prototype sequence that most others approach.

Consensus sequences are evolutionarily conserved motifs that are disclosed by multiple sequence alignments when species are compared by software algorithms for pattern recognition. Such bioinformatic analyses reveal which sequences are conserved (unchanged) over evolutionary time. Conserved sequences are often regulatory sequences that control biosynthesis or they are signal sequences that regulate maturation or direct molecules to specific intracellular sites. Evolutionary distance can be estimated by the amount of divergence of variable residues and evolutionary relatedness by the conservation of such sites.

For example, the highly conserved, consensus sequence for the 5' donor splice site is (for RNA): (A or C)AG/GUAAGU. That is, most exons end with AG and introns begin with GU (GT for DNA, image). The highly conserved, consensus sequence for the 3' acceptor splice site is (for RNA): (C/U)greater than 10N(C/U)AG/G, where most introns end in AG after a long stretch of pyrimidines. The branch site within introns (area of lariat formation close to the acceptor site during splicing) has the consensus sequence UAUAAC (image). In most cases, U can be replaced by C and A can be replaced by G. However, the penultimate (bold) A residue is fully conserved (invariant).

Left: diagram of highly conserved, consensus (DNA) sequences for 5' donor splice site, branch site, and 3' acceptor splice site (click to enlarge).

Right: diagram of percentage occurrence of (RNA) nucleotides at 5' donor splice site, branch site, and 3' acceptor splice site (click to enlarge).

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Deletion

The deletion mutation eliminates one or more nucleotides from a DNA sequence, and may produce a non-functional protein. If the number of deleted bases is not a multiple of 3, then the deletion will cause frameshift, with potentially serious consequences.


Deletion and insertion mutations often occur in repetitive sequences, such as deletion of "AT" from the sequence "ATAT" in the CFTR gene. Such mutations are most often caused by a "replication slippage", where the new strand mispairs with the template strand at repetitive sequences. Slippage can cause mispairing of several repeats. Forward slippage results in deletion mutations, while backward slippage results in insertions.


Replication slippage is mainly responsible for microsatellite polymorphisms, which are also called short tandem repeats (STR). In microsatellites, the repeat unit comprises only 1 to 6 bp and the whole repetitive region spans less than 150 bp, while minisatellites range from 1 to 20 kb, and satellites span from 100 kb to more than 1 Mb.


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External : Tandem repeats and morphological variation

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Duplication

Duplication of a segment of the arm of a chromosome is similar to insertion of sequences at the level of genes. Duplications have played an important role in the evolution of the genomes of many species.

Duplication results from unequal recombination (crossing-over) when homologous chromosomes are mis-aligned during meiosis. The frequency of duplication events depends upon the number of shared repetitive elements on the homologous chromosomes. The recombination products of duplication events include duplication at the site of exchange combined with a reciprocal deletion. Thus, duplication events produce an increase of some genetic material. Once established in a genome, functional gene duplicates persist by a variety of mechanisms.

In 1970, Susumu Ohno proposed that several rounds of genome duplication (polyploidy) occurred during vertebrate evolution (see Skrabanek and Wolfe, 1998). Ohno hypothesized that “natural selection merely modified, while redundancy created”, that is, while mutation merely provides material for possible selection, duplication creates. Ohno further hypothesized that evolutionary leaps such as the emergence of new body plans would require major events, as drastic as whole-genome duplication.[r] Study of evolutionary divergence rates in duplicated genes, and their comparison with their non-duplicated counterparts have demonstrated that "Ohno's model is correct in 95% of cases of accelerated divergence: one gene copy preserved the ancestral function, while the other copy was free to diverge. The new functions included gene silencing, anti-viral defense mechanisms, and new signaling pathways. Moreover, as a group, the ancestral and derived function genes show distinct patterns in expression, localization, and generality of function."[s]

Comparison of the tetraodon and human genomes has revealed a signature of duplicate mapping. Studying the gene correspondence between human genes and their orthologs in fish chromosomes, a team of researchers demonstrated that the human genome was doubly covered by interleaved stretches of homology to pairs of fish chromosomes, revealing that the signature of whole-genome duplication exists in the fish genome, pointing to an event common to zebrafish, fugu, and pufferfish. These results illustrate that the signatures of evolutionary change were in fact applicable across kingdoms, exhibiting basic principles of evolutionary mechanisms.[s] Comparative maps of mammalian genomes show that large chunks of chromosome are conserved between species. For example, about 200 conserved chromosomal regions (synteny) are known between mouse and human.[s]

The flowering plants, angiosperms have a marked tendency to undergo chromosomal duplication ('polyploidization') and subsequent gene loss ('diploidization').[s]

Genome sequencing suggests that the genome of yeast duplicated (became polyploid) at some point during its evolutionary past. Genome duplication in an ancestor of Saccharomyces cerevisiae occurred relatively recently, following its divergence from many other fungi such as Kluyveromyces lactis. It has been suggested that duplicated chromosomal regions in yeast are remnants of a whole-genome duplication (tetraploidy) that occurred 10^8 years ago [s]. Subsequently, many of the duplicated genes were deleted and the chromosomes were rearranged by reciprical translocation. See Yeast Gene Duplications website for details of the duplicated chromosomal regions.

Modeling gene and genome duplications in eukaryotes. modified:
Recent analysis of complete eukaryotic genome sequences has revealed that gene duplication has been rampant. Moreover, next to a continuous mode of gene duplication, in many eukaryotic organisms the complete genome has been duplicated in their evolutionary past. Such large-scale gene duplication events have been associated with important evolutionary transitions or major leaps in development and adaptive radiations of species. Here, we present an evolutionary model that simulates the duplication dynamics of genes, considering genome-wide duplication events and a continuous mode of gene duplication. Modeling the evolution of the different functional categories of genes assesses the importance of different duplication events for gene families involved in specific functions or processes. By applying our model to the Arabidopsis genome, for which there is compelling evidence for three whole-genome duplications, we show that gene loss is strikingly different for large-scale and small-scale duplication events and highly biased toward certain functional classes. We provide evidence that some categories of genes were almost exclusively expanded through large-scale gene duplication events. In particular, we show that the three whole-genome duplications in Arabidopsis have been directly responsible for >90% of the increase in transcription factors, signal transducers, and developmental genes in the last 350 million years. Our evolutionary model is widely applicable and can be used to evaluate different assumptions regarding small- or large-scale gene duplication events in eukaryotic genomes.
Maere S, De Bodt S, Raes J, Casneuf T, Van Montagu M, Kuiper M, Van de Peer Y. Modeling gene and genome duplications in eukaryotes. (Free Full Text Article) Proc Natl Acad Sci U S A. 2005 Apr 12;102(15):5454-9. Epub 2005 Mar 30.

Gene-balanced duplications, like tetraploidy, provide predictable drive to increase morphological complexity.
Controversy surrounds the apparent rising maximums of morphological complexity during eukaryotic evolution, with organisms increasing the number and nestedness of developmental areas as evidenced by morphological elaborations reflecting area boundaries. No "predictable drive" to increase this sort of complexity has been reported. Recent genetic data and theory in the general area of gene dosage effects has engendered a robust "gene balance hypothesis," with a theoretical base that makes specific predictions as to gene content changes following different types of gene duplication. Genomic data from both chordate and angiosperm genomes fit these predictions: Each type of duplication provides a one-way injection of a biased set of genes into the gene pool. Tetraploidies and balanced segments inject bias for those genes whose products are the subunits of the most complex biological machines or cascades, like transcription factors (TFs) and proteasome core proteins. Most duplicate genes are removed after tetraploidy. Genic balance is maintained by not removing those genes that are dose-sensitive, which tends to leave duplicate "functional modules" as the indirect products (spandrels) of purifying selection. Functional modules are the likely precursors of coadapted gene complexes, a unit of natural selection. The result is a predictable drive mechanism where "drive" is used rigorously, as in "meiotic drive." Rising morphological gain is expected given a supply of duplicate functional modules. All flowering plants have survived at least three large-scale duplications/diploidizations over the last 300 million years (Myr). An equivalent period of tetraploidy and body plan evolution may have ended for animals 500 million years ago (Mya). We argue that "balanced gene drive" is a sufficient explanation for the trend that the maximums of morphological complexity have gone up, and not down, in both plant and animal eukaryotic lineages.
Freeling M, Thomas BC. Gene-balanced duplications, like tetraploidy, provide predictable drive to increase morphological complexity. Genome Res. 2006 Jul;16(7):805-14.

Modeling gene and genome duplications in eukaryotes. [Proc Natl Acad Sci U S A. 2005] PMID: 15800040
Following tetraploidy in an Arabidopsis ancestor, genes were removed preferentially from one homeolog leaving clusters enriched in dose-sensitive genes. [Genome Res. 2006] PMID: 16760422
Widespread genome duplications throughout the history of flowering plants. [Genome Res. 2006] PMID: 16702410
New evidence for genome-wide duplications at the origin of vertebrates using an amphioxus gene set and completed animal genomes. [Genome Res. 2003] PMID: 12799346
Extensive genomic duplication during early chordate evolution. [Nat Genet. 2002] PMID: 12032567
See all Related Articles...

Extensive genomic duplication during early chordate evolution.
Opinions on the hypothesis that ancient genome duplications contributed to the vertebrate genome range from strong skepticism to strong credence. Previous studies concentrated on small numbers of gene families or chromosomal regions that might not have been representative of the whole genome, or used subjective methods to identify paralogous genes and regions. Here we report a systematic and objective analysis of the draft human genome sequence to identify paralogous chromosomal regions (paralogons) formed during chordate evolution and to estimate the ages of duplicate genes. We found that the human genome contains many more paralogons than would be expected by chance. Molecular clock analysis of all protein families in humans that have orthologs in the fly and nematode indicated that a burst of gene duplication activity took place in the period 350 650 Myr ago and that many of the duplicate genes formed at this time are located within paralogons. Our results support the contention that many of the gene families in vertebrates were formed or expanded by large-scale DNA duplications in an early chordate. Considering the incompleteness of the sequence data and the antiquity of the event, the results are compatible with at least one round of polyploidy.
McLysaght A, Hokamp K, Wolfe KH. Extensive genomic duplication during early chordate evolution. Nat Genet. 2002 Jun;31(2):200-4. Epub 2002 May 28. Comment in: Nat Genet. 2002 Jun;31(2):128-9.

Fugu genome analysis provides evidence for a whole-genome duplication early during the evolution of ray-finned fishes. [Mol Biol Evol. 2004] PMID: 15014147
New evidence for genome-wide duplications at the origin of vertebrates using an amphioxus gene set and completed animal genomes. [Genome Res. 2003] PMID: 12799346
Ancient large-scale genome duplications: phylogenetic and linkage analyses shed light on chordate genome evolution. [Mol Biol Evol. 1998] PMID: 9729879
Phylogenetic analyses alone are insufficient to determine whether genome duplication(s) occurred during early vertebrate evolution. [J Exp Zoolog B Mol Dev Evol. 2003] PMID: 14508816
Phylogenetic analysis of T-Box genes demonstrates the importance of amphioxus for understanding evolution of the vertebrate genome. [Genetics. 2000] PMID: 11063699
See all Related Articles...


Timing and mechanism of ancient vertebrate genome duplications -- the adventure of a hypothesis.
Complete genome doubling has long-term consequences for the genome structure and the subsequent evolution of an organism. It has been suggested that two genome duplications occurred at the origin of vertebrates (known as the 2R hypothesis). However, there has been considerable debate as to whether these were two successive duplications, or whether a single duplication occurred, followed by large-scale segmental duplications. In this article, we review and compare the evidence for the 2R duplications from vertebrate genomes with similar data from other more recent polyploids.
Panopoulou G, Poustka AJ. Timing and mechanism of ancient vertebrate genome duplications -- the adventure of a hypothesis. Trends Genet. 2005 Oct;21(10):559-67


Evolution and diversity of fish genomes. [Curr Opin Genet Dev. 2003] PMID: 14638319
Major events in the genome evolution of vertebrates: paranome age and size differ considerably between ray-finned fishes and land vertebrates. [Proc Natl Acad Sci U S A. 2004] PMID: 14757817
Fugu genome analysis provides evidence for a whole-genome duplication early during the evolution of ray-finned fishes. [Mol Biol Evol. 2004] PMID: 15014147
Genome duplication in the teleost fish Tetraodon nigroviridis reveals the early vertebrate proto-karyotype. [Nature. 2004] PMID: 15496914
See all Related Articles...

Turning the clock back on ancient genome duplication. [Curr Opin Genet Dev. 2003] PMID: 14638327
Were vertebrates octoploid? [Philos Trans R Soc Lond B Biol Sci. 2002] PMID: 12028790
Phylogenetic dating and characterization of gene duplications in vertebrates: the cartilaginous fish reference. [Mol Biol Evol. 2004] PMID: 14694077
Phylogenetic analyses alone are insufficient to determine whether genome duplication(s) occurred during early vertebrate evolution. [J Exp Zoolog B Mol Dev Evol. 2003] PMID: 14508816
Analysis of lamprey and hagfish genes reveals a complex history of gene duplications during early vertebrate evolution. [Mol Biol Evol. 2002] PMID: 12200472
See all Related Articles...

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Exaptation

The function of traits can shift under the influence of evolutionary selection, such that a trait serving one function evolves to serve another function. This process, which is commonly observed in both anatomy and behavior, was termed exaptation by Gould and Vrba.

1) A character, previously shaped by natural selection for a particular function (an adaptation), is coopted for a new use—cooptation, or
2) A character whose origin cannot be ascribed to the direct action of natural selection (a nonaptation), is coopted for a current use—cooptation.

Such shifts in the function of a trait are referred to as exaptation, cooption, or preadaptation. Feathers provide the classic example of exaptation – having initially evolved for temperature regulation, feathers later evolved for their function in flight. Human hands result from a long sequence of exaptations – fins in fish; forelimbs in transitional forms such as Tiktaalik, Acanthostega, or Ichthyostega; forelimbs in tetrapods; structures for grasping of tree branches in arboreal primates; tool-grasping structures in hominids and humans. The proteins that form the bacterial flagellum are derived from proteins that served other functions in smaller assemblages.

Interest in exaptation results from its relationship to both the process and product of evolution: to the process that creates complex traits and to the product that may be imperfectly designed.

Gould, Stephen Jay, and Elizabeth S. Vrba (1982), "Exaptation - a missing term in the science of form," Paleobiology 8 (1): 4-15.

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Frameshift

Frameshift mutations, or framing errors arise when a number other than a multiple of 3 of nucleotides are inserted into or deleted from a DNA sequence. Because a codon comprises a nucleotide triplet, indels other than multiples of 3 will cause a shift of the reading frame.

Missense mutations code for different amino acids, which may or may not alter the activity of a protein; and, nonsense mutations code for a stop codon, or premature termination codon (PTC+) which alters the code to that for a truncated protein. Nonsense mutations elicit nonsense-mediated mRNA decay (NMD).

Substitution mutations involve the replacement of one or more nucleotides with an equal number of different nucleotides – most substitutions involve a change in only one nucleotide (point mutations, single nucleotide polymorphisms). Because an equal number of nucleotides are substituted, frameshift is not a problem.

Deletions or insertions of short regions can occur by strand slippage, and deletions or insertions of longer regions can occur via homologous recombination. Deletion and insertion mutations often occur in repetitive sequences, such as deletion of "AT" from the sequence "ATAT" in the CFTR gene. Such mutations are most often caused by a "replication slippage", where the new strand mispairs with the template strand at repetitive sequences. Slippage can cause mispairing of several repeats. Forward slippage results in deletion mutations, while backward slippage results in insertions.

Replication slippage is mainly responsible for microsatellite polymorphisms, which are also called short tandem repeats (STR). In microsatellites, the repeat unit comprises only 1 to 6 bp and the whole repetitive region spans less than 150 bp, while minisatellites range from 1 to 20 kb, and satellites span from 100 kb to more than 1 Mb.

Mobile elements called insertion sequences exist in nature. These sequences encode only the information necessary for their insertion into DNA. Depending upon the particular insertion sequence, they can insert at specific regions or at random.

If the number of inserted bases is not a multiple of 3, insertion will cause frameshift, with serious consequences. A number of diseases are caused by insertions without frameshift - Huntington's chorea, Myotonic Dystrophy, Fragile X site A, Fragile X site E, Fragile X site F, Kennedy disease, SCA1, DRPLA.

External : Tandem repeats and morphological variation

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Insertion

Mutations develop from both insertion and loss of segments of DNA. Deletions or insertions of short regions can occur by strand slippage, and deletions or insertions of longer regions can occur via homologous recombination.


Diagram of insertion of a segment of chromosome a into chromosome b, resulting in shortened chromosome a' and lengthened chromosome b' (click to enlarge image).

If the number of inserted bases is not a multiple of 3, insertion will cause frameshift, with serious consequences. A number of diseases are caused by insertions without frameshift - Huntington's chorea, Myotonic Dystrophy, Fragile X site A, Fragile X site E, Fragile X site F, Kennedy disease, SCA1, DRPLA.

Proteins gradually evolve by the accumulation of mutations.

Mobile elements called insertion sequences exist in nature. These sequences encode only the information necessary for their insertion into DNA. Depending upon the particular insertion sequence, they can insert at specific regions or at random.

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It has been known since the beginning of the twentieth century that unstable or variable gene loci occur in plants. Breakage-fusion (reunion)-bridges are formed during anaphase whenever two chromosomes fuse at their ends, generating a fusion product with two centromers. If these two fused chromosomes are subsequently carried to different poles than the regular chromosomes, a chromosomal fraction results. During the subsequent S-phase, the chromatid with a fused-fraction at its terminus will replicate, leading again to a fusion of the homologous chromatids. Consequently a chromosome comprising one chromatid with two centromeres will occur in the subsequent mitosis, rather than a chromosome from two chromatids and one centromere. Consequently, a second fraction occurs during anaphase when the second round of the cycle starts.

B. McClintock recognized (between ‘47 and ‘51) that the chromosomal fraction is restricted to certain sections of the chromosome, which she termed Ds (dissociation). The Ds segment is a mutator gene, which behaves like a pseudoallele that can be located at different gene loci. This mutator gene can insert itself into other genes, rendering them inactive. Thus, it is a control element that changes its location within the chromosome, causing mutations wherever it inserts. Such mutator genes are also called "jumping genes".

A further set of elements, the Ac (activation) elements, support the chromosomal fraction or a translocation of a Ds element. An Ac element can be regarded as a multiple allele, and it may occur different sites in all chromosomes. A number of gene loci are known to be influenced by the Ds-Ac-system or other control elements. Detection of the spm-system (suppressor-mutator) and the elucidation of its function established that the control elements not only act as switches (a yes/ no decision) but that they also modulate the degree of gene expression.

Insertion elements and transposons were first detected in bacterial DNA during the late sixties. This discovery explained the connection between transposons and the chromosome fraction control elements.

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External : Transposons part 1, transposons part 2 : Barbara McClintock and mobile genetic elements :

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Inversion

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Meiosis

Meiosis produces germ cells, or gametes with half (haploid) the number of chromosomes of a diploid cell.

First cell division: meiosis I: single diploid parent cell successively passes through:
1. prophase 1
2. metaphase 1
3. anaphase 1
4. telophase 1 & prophase 2

Second cell division: meiosis II:
5. metaphase 2
6. anaphase 2
7. telophase 2,
producing 4 haploid daughter cells.

Recombination provides a mechanism for genetic variability and is a mechanism of biological evolution. Recombination between homologous chromosomes in meiosis I involves the formation and repair of double strand breaks (DSBs), and meiosis I employs the same enzymes as does DSB repair. Many biologists consider the main function of sexual reproduction is to provide this mechanism for maintaining the integrity of the genome.

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mispairing

Mispairing refers to the presence of at least one nucleotide in one strand of a DNA molecule, which is not the complement of the nucleotide at the corresponding locus in the other strand.

Slipped strand mispairing occurs during DNA replication. Regions of DNA that are capable of assuming hairpin-like secondary structures are particularly prone to this error because displacement of the strands disrupts alignment of bases. Mispairing results in the repeated replication of the same stretch of DNA, and provides an explanation for satellite DNA.

Deletion and insertion mutations, indels, often occur in repetitive sequences. For example, deletion of "AT" from the sequence "ATAT" in the CFTR gene. Such mutations are most often caused by a "replication slippage", where the new strand mispairs with the template strand at repetitive sequences.

Tandem repeat sequences are particularly prone to high mutation rates because of mispairing and replication slippage. Slippage can cause mispairing of several repeats. Forward slippage results in deletion mutations, while backward slippage results in insertions. Replication slippage results in mispairing not because the nucleotides are altered, but because the number of repeats may vary between one strand and the other. Such repeats are retained in sections of the genome that can tolerate variability.

Replication slippage is mainly responsible for microsatellite polymorphisms, which are also called short tandem repeats (STR) or simple sequence repeats (SSR). In microsatellites, the repeat unit comprises only 1 to 6 bp and the whole repetitive region spans less than 150 bp, while minisatellites range from 1 to 20 kb, and satellites span from 100 kb to more than 1 Mb. SSRs occur throughout the genome, while tandem repeats occur in telomeres and centromeres. The nucleotide sequence of repeats is fairly well conserved across a species, but variation in the length of the repeat is common, as in VNTRs, or variable number tandem repeats.

Once an alteration (such as a point-mutation) occurs within a tandem repeat, the cell's replication machinery is able align the two DNA strands and the alteration will be replicated and inherited. Subsequent deletion of the alignment-promoting alteration would restore slippage. Thus, purity of repetition indicates a lengthy history of deletion and expansion. Strands of repeated sequences can code for polypeptides with repeated amino acids, and the ratio of such poly-amino-acid chains can regulate transcription factors. Polyglutamine can increase the rate of transcription, while polyalanine reduces it. Polyadenylation is a stage of RNA processing in which the 3’ end of the pre-mRNA is cleaved before a stretch of adenosines are added to the end of the molecule.

Animation of slipped strand DNA mispairing DNA Animation :
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External : Tandem repeats and morphological variation

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Recombination

Recombination occurs during meiosis and involves a shuffling of genes, resulting in genetic variation.

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Substitution

Substitution mutations involve the replacement of one or more nucleotides with an equal number of different nucleotides – most substitutions involve a change in only one nucleotide (point mutations, single nucleotide polymorphisms).

Transition mutations involve the substitution of a pyrimidine nucleotide for a pyrimidine, or a purine nucleotide for a purine. Transversion mutations involve switching between a pyrimidine nucleotide and a purine.

Substitution mutations generate silent, missense, or nonsense mutations depending upon the consequences of the substitutions. Silent mutations code for the same amino acid in the protein; missense for different amino acids, which may or may not alter the activity of a protein; and, nonsense for a stop codon, which alters the code to that for a truncated protein. Accordingly, substitution mutations can be neutral, beneficial, or deleterious.

Missense point mutations can transform the Ras proto-oncogene into the Ras oncogene. Mutations of Ras proto-oncogenes are common → H-RAS, N-RAS, and K-RAS oncogenes. Inappropriate activation of the Ras gene plays a key role in signal transduction, proliferation, and malignant transformation. Mutagenic mutation alters the Ras protein so that the GTPase reaction can no longer be stimulated by GAP, increasing the half life of active Ras-GTP mutants and ensuring that this molecular switch remains 'on'. Unchecked proliferation occurs because mutations that prevent GTP hydrolysis favor constitutive activation as RAS-GTP. The commonest neoplastic mutations are at the 12 (G to T transversion, Gly→Val) → GAP insensitive, and the 61 positions → stabilizing against GTP hydrolysis.
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Translocation

Translocation involves the exchange of sections of genetic material between non-homologous chromosomes.

In balanced translocations, the total complement of a cell's genetic material is normal, with material merely located on non-homologous chromosomes. In unbalanced translocations, which are associated with disease, cells contain an increased or reduced complement of genetic material. Translocations are also classified as Robertsonian (resulting from fusion of acrocentric chromosomes) or reciprocal (resulting from exchange of segments of chromosomal arms).

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