Course Name : BSC Subject Name

1 Course Name : BSC Subject Name: Plant Biotechnology Prepared by Assistant Professor’s Team of Microtek College of Management & Technology Under Guidance of Dr. Pankaj Rajhans An Alumni of IIT-Delhi President & Executive Director Microtek College of Management & Technology Jaunpur & Varanasi (U.P) 2 Unit I Plant genome organization (Page 3-8) Structure of representative plant genes Gene families in plants Organisation of chloroplast genome Nuclear encoded and chloroplast encoded genes for chloroplast proteins Targeting of proteins to chloroplast Organisation of mitochondrial genome - encoded genes for mitochondrial proteins RNA editing for plant mitochondria Cytoplasmic male sterility Seed storage proteins Maize transposable elements, their organisation and function Transposabie elements in transgenic plants Regulation of gene expression in plant development -Plant hormones and phytochrome Unit II Symbiotic nitrogen fixation in legumes by Rhizobia Nitrogen fixation in Cyanophyta the biochemistry molecular biology and gene rearrangement Agrobacterium and crown gall tumors Mechanism of TDNA transfer to plants Ti plasmid vectors and its utility Classification and molecular biology of plant viruses Molecular biology of -plant stress response Transgenic plants and applications vaccine and other biological developments Unit III Genetic engineering in plants Selectable' markers Reporter genes and promoter used in plants by physical means Genetic engineering of plants for construction of genome libraries and CDNA libraries Molecular breeding 3 PLANT BIOTECHNOLOGY Plant Genome - Genetically, a plant genome is the most complex one found in living systems. It comprises three interacting genome. Aside from the nuclear genome, complete genetic systems are located in the plastids and the mitochondria. These organelles are semiautonomous bodies; they have their own organizational and functional properties but do not synthesize all their own proteins. The nuclear genome plays an important role in organelle biogenesis. Techniques in molecular cloning and deoxyribonucleic acid (DNA) sequencing have provided the tools for studying the structure of genes at the nucleotide level. Hence our knowledge of the structure, organization and expression of a plant genome has come largely through, the use of recombinant DNA techniques. This technology allows isolation and characterization of specific pieces of DNA by cloning the DNA sequences into bacterial cells in which they can be replicated and large quantities obtained for analysis. In addition to supplying much basic information concerning gene structure and expression, recombinant DNA technology enables specific manipulation of genetic material and the transference of such material between different organisms. These types of genetic manipulations may ultimately give us the opportunity to engineer agricultural crops and industrially important plants in order to, tailor them more closely to fulfill the needs of human beings. Nuclear Genome - A genome is the complete set of chromosomes found in each nucleus of a given species which contains the entire genetic material. The nuclear genome is the largest in the plant cell, both in terms of picograms of DNA and in number of genes encoded (complexity). Eukaryotic nuclear genomes can be distinguished from organelle and prokaryotic genomes by size and complexity. A typical higher plant genome, for example, contains about 5 x 109 base pairs of DNA per haploid set of chromosomes. This is about 30,000 times as much DNA as in a single chloroplast genome and some 10,000 times as much as in a moderately sized plant mitochondrial genome. It is also 1000 times more than that of bacterial DNA present in Escherichia coli. The typical plant genome with 5 x 109 base pairs of DNA would be about three metres long if the entire DNA were to be laid out in a straight line. Chromosomes are composed of two types of large organic molecules (macromolecules) called proteins and nucleic acids. Nucleic acids are of two types: deoxyribonucleic acids. (DNA) and ribonucleic acids (RNA).For many years scientists disputed which of these three macromolecules (proteins, RNA and DNA) carries genetic information. 4 During the 1940s and early 1950s several elegant experiments were carried out which clearly established that genetic information resides in the nucleic acids and not in the proteins. More specifically, these experiments showed that genetic information resides in DNA. (In a few simple viruses, however, RNA carries the genetic information; these particular viruses contain no DNA.) Nuclear DNA is packaged into chromosomes along with histones and nonhistone proteins, all of which play important roles in gene expression. These various components are held together to form chromatin by both hydrophobic and electrostatic forces. While the DNA encodes the genetic information, the proteins are involved in controlling packaging of DNA and in regulating its availability for transcription. Although the structure of eukaryotic chromatin has been fairly well characterized, the roles of various individual proteins in chromatin structure and gene regulation have yet to be elucidated. Size of Nuclear Genome of Various Plant Species – Species pg DNA per Haploid Genome Arabidopsis thaliana 0.50 Vigna radiata 0.53 Nicotiana tabaccum 2.00 Phaseolus coccineus 3.50 Triticum aestivum 5.10 Secale cereale 8.40 Vicia faba 26.00 Allium cepa 33.50 Viscum album 107.00 Repeated Sequences of Regulating Gene Expression - In a strict sense, some of the control sequences just described is really repeated sequences. Short elements, such as the TATA or CAAT boxes, appear in many promoters and hence are repeated, but because they are so short are not recognized in hybridization experiments. Enhancers may sometimes contain several repeats of a somewhat longer sequence (such as the 72-nucleotide repeats of the SV40 enhancer), By themselves, however, these do not reach sufficiently high copy numbers to be detected in classical reassociation experiments and no extensive sequence homology exists among enhancers as a class, although some short homologies have been found. Therefore, none of the gene regulatory elements discussed thus far can be considered "classical" repeated sequences. 5 Evidence for the involvement of "classical" repeated sequences in gene regulation is far more circumstantial and is based on the occurrence of repeat sequence transcripts in the RNA population. Although most mRNAs are transcripts of single copy genes, repeat transcripts can often be found when Cloned repeat sequence probes are used to increase sensitivity. This is especially true in nuclear RNAs, wherein the RNA contains transcripts of many interspersed repeat sequences. When different stages of develop ment are compared, it is usually found that the repeat sequences which are most abundant in RNA differ from stage to stage. Of course, simply because repetitive sequences are transcribed, or because their transcription changes during development, does not prove that they are involved in gene regulation. It seems likely that most transcribed repeats are simply sequence elements that have found their way into transcription units by evolutionary accidents or rearrangement or transposition. On the other hand, if must be admitted that a small class of functionally important repeat sequences might exist against a large background of evolutionary noise. Several lines of evidence from animal systems suggest that this may be true. The transcriptional elements are parts of a gene or its flanking sequences and affect only the gene to which they are directly linked. In genetic terms they function cis, regulating genes on the same chromosome. Clearly, there must be other elements involved in gene regulation and, as a matter of fact, there is now good experimental evidence that in some cases certain protein factors must be bound to specific sites in a gene or its flanking sequences in order to initiate transcription. An example is the transcription factor which binds to the heat shock promoter in Drosophila. Genes encoding this factor would be examples of trans-acting control elements. As their name implies, such elements are capable of affecting genes other than those located on the same chromosome. Transacting regulatory genes are well known from genetic studies but our understanding of the molecular nature of such elements, and the factors which presumably mediate their effects, is still quite limited. Certainly, their importance can hardly be denied. Gene families:- This is a list of gene families or gene complexes, that is sets of genes which occur across a number of different species which often serve similar biological functions. These gene families typically encode functionally related proteins, and sometimes the term gene families is a shorthand for the sets of proteins that the genes encode. They may or may not be physically adjacent on the same chromosome. Example of these:- CalcinPEst *calcineurin-like phosphoesterase 6 Calmod Calmodulin CytP450 *cytochrome P450 Enod16 Enod16 Protein family:-A protein family is a group of evolutionarily related proteins, and is often nearly synonymous with gene family. The term protein family should not be confused with family as it is used in taxonomy. Proteins in a family descend from a common ancestor (see homology) and typically have similar three-dimensional structures, functions, and significant sequence similarity. While it is difficult to evaluate the significance of functional or structural

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