Pengantar Bioteknologi

Bioteknologi :Teknologi yang menggunakan sistem hayati (proses-proses biologi) untuk mendapatkan barang dan jasa yang berguna bagi kesejahteraan manusiaIlmu dasarnya : mikrobiologi, genetika molekuler, biosel, biokimia, bio molekuler, proses rekayasaProses: fermentasi (steril, non steril), cangkok (gen, inti), rekombinasi genetik, hibridoma, kultur jaringan,

Pengertian

SejarahMinuman alkohol anggur(pengawetan daging)

Khamir rotiKejuYogurtSusu masamkecap1857 Pasteur menemukan bahwa fermentasi merupakan proses yang dilakukan oleh organisme hidup1920 fermentasi aseton, etanol, butanol, gliserin

PD II Penicillium natatum Antibiotik Vitamin, steroid, enzim

1859Charles Darwin publishes his book (On the Origin of Species by Means of Natural Selection). According to this book evolution is lifes motor. The interplay of mutation and selection endows living beings with optimized traits in order to survive. These principles are also valid for the so-called 'chemical evolution' of biomolecules and are being used in laboratories for in vitro optimizing of wanted qualities in molecules

1865Gregor Mendel finds that independent 'factors' are responsible for the heredity of traits from one generation to the next according to a set of (Mendelian) priciples.

1869Friedrich Miescher discovers an acidic substance in the nucleus of cells which he names 'nuclein'. By elemental analysis he finds 14 % nitrogen, 3 % phosphorus, and 2 % sulfur (from proteins). Since the substance cannot be cleaved by the proteolytic enzyme pepsin Miescher concludes that this substance is not a protein.

1879Walther Flemming observes the separation of chromosomes during mitosis but fails to fully understand its meaning. He counts 24 pairs of chromosomes, a number which will be corrected to 23 later in 1956 by the Indonesian scientist Joe-Han Tijo.

1900William Bateson introduces 'genetics' as a scientific disciplineHugo de Vries, Erich Tschermak von Seysenegg and Carl Correns independently rediscover the Mendelian principlesHugo de Vries defines the meaning of 'mutations'

1902Walter Sutton observes in grasshoppers' cells that chromosomes carry the Mendelian 'factors of heredity', i.e. the genetic information.

1909Wilhelm Johansen coins the terms 'gene', 'genotype', and 'phenotype' referring to Mendel's 'factors'. Archibald Garrod confirms the hereditary nature of four metabolic diseases. He publishes "Inborn Errors of Metabolism"

1910Thomas Hunt Morgan's studies in the biology of Drosophila melanogaster confirm that certain traits are inherited sex-specifically. He also proves that some phenotypes result from several genes located on different chromosomes.

1927Herman Muller finds that energetic radiation causes defects in the chromosomes, i.e. mutations.

1928Fred Griffith demonstrates that material from killed bacteria of the pathogenic strain Streptococcus pneumoniae S ('smooth') is taken up from living ones of the non-pathogenic strain Streptococcus pneumoniae R ('rough') which are subsequently 'transformed' into the pathogenic S-strain. Pneumococcus pneumoniae

1944Oswald Theodore Avery, Colin McLeod, and Maclyn McCarty find by careful analysis of Griffith' experiments that desoxynucleic acid (DNA) is the carrier of the 'transforming principle'.

1945Erwin Schrdinger proposes in his famous book 'What is life?' that genes must be 'aperiodic crystals' consisting of a succession of a small number of isomeric elements whose precise sequence constitutes the heredity code. Although these ideas do nothing to identify the responsible molecular structures, they attract many newcomers to the field.

1950Erwin Chargaff observes the 1:1 ratio of the nucleic bases adenine/thymine and guanine/cytosine. This will be a decisive hint for Watson & Crick to the structure of the DNA molecule.

1952Alfred Day Hershey's and Martha Chase's studies in bacteriophages prove that only the nucleic acids carry the genetic information; proteins are definitively excluded from playing this role. Joshua Lederberg finds plasmids in bacteria, small rings of extrachromosomal DNA which can be replicated autonomously.

1953James D. Watson and Francis H. C. Crick elucidate the structure of the DNA molecule. It consists of two strands which are bound by hydrogen bonds bewtween opposite basepairs of adenine-thymine and cytosine-guanine. Their model is based on the results of many colleages, namely Rosalind Franklin who discovered the helical structure and the outward position of the phosphate-sugar backbone.

1955Frederick Sanger finds a method to sequence proteins. With this he determines the sequence of the amino acids of insulin.

1956..Arthur Kornberg reports the first in vitro synthesis of a DNA molecule. George Emile Palade locates protein biosynthesis to the ribosomes, the 'factories of the cell'.

1956Joe Han Tijo and Albert Levan correct the number of the chromosome pairs in human cells to 23. Since the days of Walther Flemming 24 was believed to be the true number. Chromosomen einer menschlichen Zelle protein chains are built at the ribosomes

1959T. Akiba, T. Koyama, Y. Isshiki, S. Kimua, T. Fukushima, T. Watanabe and T. Fukusawa describe the transfer of multiple resistance to antibiotics by bacterial plasmids. Plasmids turn out to be efficient vectors for trafficking genetic information between bacteria of different strains. bacterial plasmid (arrow)

1960Francois Jacob and Jaques Monod recognize the function of the messenger RNA (mRNA). They develop the operon model of gene regulation in procaryotes.

1961Sydney Brenner and Francis Crick claim that every amino acid corresponds to a triplett of nucleotides, called 'codons'. Marshall Nirenberg and Heinrich Mathaei can prove that the codon UUU in mRNA codes for the amino acid phenyl alanine Sydney BrennerMarshall Nirenberg

1962John Gurdon claims to have reproduced frogs from the epithelium cells of Xenopus laevis. J. B. S. Haldane coins 'cloning' when describing these debated experiments.Xenopus laevis South African clawed frogs John Gurdon

1966Mainly due to the work of Har Gobind Khorana the 'Genetic Code' is completely known.

1968Werner Arber discovers enzymes, so-called nucleases, which digest DNA double strands from the ends.

1969Jonathan Beckwith is the first to isolate a complete gene, in this case a gene from the bacterial sugar metabolism.

1970Hamilton Smith discovers an enzyme (HindII) which specifically cleaves DNA double strands Har Gobind Khorana synthesizes a complete gene (that of an alanine t-RNA) in vitro. David Baltimore and Howard Temin discover the viral enzyme reverse transcriptase. It translates RNA sequences to DNA sequences and thus tumbles the age old 'Central Dogma of Genetics' which defined the direction of the flow of the genetic information to start from DNA via RNA to the proteins.

1971Paul Berg, Peter Loban, and Dale Kaiser find enzymes which attach short single strands to the blunt ends of double stranded DNA. The single strands consist of only one sort of nucleotides. Complementary single strands (sticky ends) allow the coupling of ds DNA fragments; the remaining gaps can be closed by the use of ligases.

1972Janet Mertz and Ron Davis paste DNA fragments of different origin resulting from specific enzymatic restriction. They close the gaps using ligases and thus produce recombinant DNA.

1973Stanley Cohen and Herbert Boyer paste enzymatically restricted DNA fragments into plasmids which have been cleaved by restriction enzymes. The resulting recombinant plasmids can serve as vectors for the transfer of foreign DNA into bacteria. These experiments are generally regarded as the beginning of the era of genetic engineering and modern biotechnology.

1975The first conference on safety issues of the new technology is held at Asilomar in California.Georges Khler and Cesar Milstein develop the 'Hybridoma Technology' for the production of monoclonal antibodies. An antibody producing plasma cell (derived from a B lymphocyte) is fused with a tumor cell resulting in an immortal hybridoma cell which (as well as its daughters) continues to produce one kind of antibodies.

1976Herbert Boyer and Robert Swanson found Genentech the first biotech company.

1977Walter Gilbert and Frederick Sanger independently develop two different methods for DNA sequencing.

1978Walter Gilbert discovers that genes of eucaryotic organisms are composed of coding and non-coding parts. The non-coding sequences, named introns, are cut from the mRNA before translation leaving the coding exons as the source of information for protein synthesis.

1979Thomas Cech and Sydney Altman discover the autocatalytic activity of some RNA molecules to cleave themselves at well defined positions. They coin 'ribozyme' for this type of RNA.

1980Jozef Schell and Marc van Montagu transfer foreign DNA into plant cells by employing T-plasmids of Agrobacterium tumefaciens as vectors.

1981The first patent for a genetically modified organism is granted to Ananda Chakrabarty of General Electric. It covers a P. aeruginosa strain which is equipped with genes of certain enzymes in order to metabolize crude oil.

1982The U.S. Food and Drug Agency (FDA) approves recombinant insulin for marketing.

1984Alec Jeffreys develops 'genetic fingerprinting' allowing the comparison of minute variations, the so-called restriction length fragment polymorphisms (RLFPs), in the genomes of two organsims.

1985Karry Mullis develops the polymerase chain reaction (PCR). This powerful tool allows to copy and accumulate extremely low amounts of DNA from various sources until a level sufficient for analysis is reached.

1986Neal First develops protocols for clonig cattle from bovine embryos by separating cells at early embryonic stages. These experiments resemble John Gurdons earlier attempts at the cloning of frogs. The first cloning of an adult animal will be reported in 1997 by Ian Wilmut who succeeds in cloning a sheep, Dolly.

1988The first patent for a transgenic mammal is granted to Harvard's Philip Leder and Timothy Stewart. It refers to a mouse which is susceptible to tumors and serves as model organism for studies in cancer.

1990French Anderson successfully tries the first somatic gene therapy of a human. Patient Ashanti DeSilva is cured from an deficiency in the enzyme ADA, which causes severe immunodeficiency. German 'Genetic Engineering Act' passes parliament Official kick-off of the Human Genome Project (HGP)

1993The German Genetic Engineering Act of 1990 is revised, i.e. many restrictions of the 1990 version are eased. The Biotechnology Industry Organization BIO is created by merging two smaller trade associations

1994Genetically modified tomatoes are on sale in the U.S. Cans containing puree from transgenic tomatoes are on the shelves of british supermarkets.

1995The Institute for Genomic Research (TIGR) publishes the first complete sequence of the genome of a free-living organism, the bacterium Haemophilus influenzae Germany launches the BioRegio contest. 17 regions compete in presenting the best blueprint for the development of commercial biotechnology to an international jury. The three winning regions will share the price of DM 150m.

1996The complete sequencing of the baker's yeast' genome is accomplished by close collaboration of laboratories in Europe, Japan, and the U.S. The Saccharomyces cervisiae genome ist the first eucaryotic genome completely sequenced. Patrick Browne of Stanford University presents the first 'gene chip' containing 6116 different gene specific sequences of the baker's yeast genome. In the U.S. transgenic plants grow on more than 1.9 million hectares

1997Ian Wilmut (right) visits Dolly (center) the first mammal cloned from an adult cell.

1998Two research teams report embryonic stem cells differentiate to specialized tissue cells. The picture shows an early developmental stage, the blastocyst. The first genome of a multicellular organism (Caenorhabditis elegans) is sequenced.Craig Mello und Andrew Fire find that small double-stranded RNA molecules can selectively block gene expression in Caenorhabditis elegans, a phenomenon later referred to as RNA interference (RNAi)

1999Pluripotent stem cells from tissues can be reprogrammed to other cell types. The sequence of chromosome 22 is published. The U.S. biotech company Celera starts its own human genome sequencing project

2000There are 1300 biotech companies on each side of the Atlantic. About 400 are listed at stock exchanges. The genome of Drosophila melanogaster is sequenced. The fruit fly was introduced by Thomas H. Morgan into genetics research. The Human Genome Organization and Celera present the first working draft of the human genome. The picture shows Celera's CEO J. Craig Venter (left) and Francis Collins, the speaker of the HGP.

2001Less genes than expected The international Human Genome Project (HGP) and biotech company Celera Genomics publish their versions of the sequence of the human genome. The Celera team estimates the number of genes to range from 26,000 to 39,000, HGP scientist' estimate is between 30,000 and 40,000. Rice genome Syngenta (Basel, Switzerland), in collaboration with Myriad Genetics (Salt Lake City, UT), announced the completed sequencing of the genome of Oryza sativa japonica

2001RNA interference Thomas Tuschl is the first to demonstrate that RNA interference (RNAi) is also working with mammalian cells.

2002Genomes of the malaria agent and its carrier sequenced Researchers of the Institute for Genomic Research (TIGR), the Sanger Institute (UK), and Stanford University publish the genomic sequence of Plasmodium falciparum and Plasmodium yoelii. The protozoa P. falciparum causes malaria. The genome of its carrier Anopheles gambiae is also sequenced and published.

2003Genome of SARS virus sequenced Within three weeks after its discovery scientists of the Michael Smith Genome Sciences Centre in British Columbia (Vancouver, Canada) succeeded in sequencing the genome of the virus causing severe acute respiratory syndrome (SARS). The RNA virus remotely resembles corona virus species and contains roughly 30,000 genes.

2003Germ cells from stem cells Karin Hbner and Hans Schler of the University of Pennnsylvania succeeded in generating oocytes from murine stem cells which were able to grow into further embryonic stages. The experiment demonstrates the potential of pluripotent stem cells to acquire totipotency via a germ line cell stage

200499.999% accuracy Three years after the publication of the first draft a far more precise version of the sequence of the human genome is published. The number of identified genes decreases again compared with previous estimations. 19,599 have been confirmed to encode proteins, 2188 are suspected to encode for proteins. 1183 genes have been generated through duplication of genes, 33 genes appear to have been degenerated into disfunctional pseudogenes.

2004Fatherless mouse Tomohiro Kono of the Tokyo University of Agriculture presented the first mouse which was obtained from oocytes only. His team combined the genomes of oocytesfrom adult and newborn mice and eventually one of 460 experiments resulted in a living animal, being 14 month old at the time of publication. Critical for success was the suppression of a gene which normally controls the imprinting of chromosomal DNA from mother and father.

2004Mice from cancer cells Rudolph Jaenisch of MIT and Lynda Chin of the Dana Farber Cancer Institute generated cloned mice from cancer cells by stuffing the nuclei of melanoma cells into de-nucleated mouse oocytes. The resulting blastocysts yielded stem cells which were suitable for transplantation into normal blastocysts which eventually developed into living animals. Some genetic markers of the melanoma cells were found again in the animal, but the epigentic patterns had gone lost - showing that epigenetic modifcations are both reversible and late stage in the transformation of normal cells into melanoma cells.

Bioteknologi tradisional :Seleksi bahan, mikrobia yang digunakan dan modifikasi lingkungan untuk memperoleh produk optimal. Misal : pembuatan tempe, tape, roti, pengomposan sampahBioteknologi modern :Memanfaatkan ketrampilan manusia dalam melakukan manipulasi makhluk hidup agar dapat digunakan untuk menghasilkan suatu barang yang diinginkan. Misal rekayasa genetikPengertian & Sejarah

Bioteknologi merupakan penerapan prinsip ilmiah dan rekayasa pengolahan bahan oleh agen biologi untuk menyediakan barang dan jasaRekayasa genetik :Teknik untuk menghasilkan molekul DNA yang berisi gen baru yang diinginkan atau kombinasi gen-gen baru atau dapat dikatakan sebagai manipulasi organisme

Hasil-hasil rekayasa genetika tanaman yang digambarkan dalam National Enquier, c. 1981.

Kesetabilan gen asing atau dapat berubah dan diekspresikan berbeda oleh organisme yang tertular gen asing mutasiPenarikan gen asing jika masuk dalam organisme non targetTerganggunya rantai makanan'Terganggunya organisme non target

2) Tinjauan kesehatanGangguan kesehatan karena produk gen asing (dalam waktu panjang)Bahaya atau tidaknya gen-gen penyerta

Tinjauan sosial dan budayaEtika, moralAgama

Perdagangan global

Aplikasi bioteknologi ?Aplikasi farmasi : antibiotik, vaksin, bahan diagnostik (enzim, antibodi), steroid, hormon, interferon, vitamin, asam amino.makanan dan minuman: produk susu (kiju, yoghurt), kopi, minuman beralkohol, pengembang roti, citarasa, sirup

bahan kimia: enzim, polimer, alkohol, aceton, butanol, asam organik (as. sitrat, as. malat ..), peluluh logam besi, tembaga, emas dll.

bahan energi: ethanol (gasohol), methane (biogas)

aplikasi pertanian : makanan ternak, vaksin ternak, pengomposan, biopestisida, fiksasi nitrogen, kultur sel dan jaringan, tumbuhan transgenik, varietas tumbuhan barulingkungan: pembersihan air, pengelolaan limbah, pembersihan limbah minyak

A. Fermentasi non sterilBahan: selulosa, etanol, minyak bumi, kertas, limbah industri / kehutananBakteri pelaku: berbagai bakteri, spirulina, sacharomices, candida utilis, methylophilusProduk: PST untuk ternak (silase), kompos, pembersihan limbah, biogas dll.

B. Fermentasi sterilproduk susu (dairy). dg. enzim renin, dibentuk casein/keju : romano/parmesan- sangat keras (an.) cheddar/swiss--- keras (an.) requifort/biru --- semi lunak (aerob) camembert --- lunak (aerob)dengan Strep. thermophilus & Lact. bulgaricus dibuat yoghurt, kefir dan kumiss

C. Fermentasi sterilproduk non susu bahan dari berbagai karbohidrat dengan Sacharomices, Asp. orizae dan Asp. wentii dibuat adonan roti, tuac, acar, olive dan kecapbahan dari serelia, kentang, anggur dan tebu dengan Sacharomices dibuat alkohol (wiski, vodka, rum)

` D. Microbiologi industriDengan Corynobacterium glutamicum dihasilkan AA lisin, treonin dan MSGDari gula tetes/sirup dengan Asp. niger dihasilkan as. sitratDengan Pseudomonas, propioni bacterium dan Ashbya gosipii dihasilkan vitamin B-12 dan riboplavinProduksi penicilin, sefalosporin,vaksin, streptomisin, interferon, antibodi,

pertahanan anti virus &anti bacteri

Reseptor antigen thd. virus/bacteri

Antigen dilawan macrofag / antibodi

T-cell clonal proliferation

D. Microbiologi industriBacteri minus-es, anti pembentukan esRhizobium dan Azotobacter untuk fiksasi nitrogenBatuan yg. mengandung Cu SO4 dan Fe(SO4)3 dioksidasi dengan H2SO4 dan Thiobasilus ferooksidan menghasilkan CuSO4 dan FeSO4 yang murni

KOREKSI GENETIK

Terapi genetic

PERKEMBANGAN KANKER

Rekayasa genetika

E. Rekombinasi genetikaYg. alami dengan cross. over pada meiosis dan mutasiYg. buatan/invitro dg. rekayasa genetika/ cangkok gen/ transplantasi genisolasi gen --- proliferasi gen dg. PCR --- insersi/cangkok gen pada bacteri/plasmid --- reflikasi, proliferasi sel --- hasilnya: produksi zat (metabolit, hormon, enzim, antibodi ), tahan thd. sifat tertentu, tanaman transgenik

TRIPLETBASANITROGEN

F. Hibridoma, cangkok intiHibridoma = fusi 2 sel, misal sel pancreas(insulin) + sel kanker = hasilnya peningkatan pada jumlah sel, antibodi, hormon, interferon inti sel terdiferensiasi pada ovum yang non diferensiasi dihasilkan cloning vegetatif

Kerja enzim retriksi

Reflikasi plasmid

Reflikasi plasmid ganda

G. Kultur Jaringan bertujuan untuk micropropagasi dan produksi metabolit scunderseleksi bibit --- pemisahan jaringan --- sterilisasi jaringan dg. blecing dan detergent --- cuci dg. air steril --- simpan pada medium tumbuh --- transfer pada medium multiplikasi --- medium pembentukan pucuk/akar/ embrio --- transfer pada tanah steril.

Perbungaan yang cuma 6 bulan dibanding normalnya 20 tahun

Trisomi-21, sindrom Down

Siklemia oleh gen resesif

NUKLEOTIDA

Terima kasih

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