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“It is one of the most beautiful compensations of this life that no man can sincerely try to help another without helping himself” -Ralph Waldo Emerson Transcription and Translation Chapters 13, 14 and 15 DNA Structure DNA Structure DNA is comprised of nucleotides (adenine, cytosine, guanine, thymine) that are connected by a sugar (deoxyribose) phosphate backbone. Phosphate group Nitrogenous Bases Sugar deoxyribose adenine (A) thymine (T) Hydrogen bonds guanine (G) cytosine (C) DNA Replication 1. DNA to be replicated 2. Strands separate 3. Each strand now serves as a template for the synthesis of a separate DNA molecule as free nucleotides base-pair with complementary nucleotides on the existing strands. Order of bases encodes Information for protein production. 4. This results in two identical strands of DNA. DNA Replication DNA Replication Review Differences between DNA and RNA Deoxyribonucleic acid (DNA): primary information bearing molecule in living things. Codes for proteins De-oxy = one less ________ on sugar (ribose) Ribonucleic acid (RNA): transfers genetic code from DNA in the nucleus to proteinforming system in the cytoplasm Uracil instead of Thymine Differences between DNA and RNA RNA is single stranded and contains _______ instead of thymine DNA is double stranded (double helix) and contains ________ instead of uracil Protein Synthesis: Transcription and Translation Transcription: process by which a gene’s base sequence is ______ to mRNA Translation: process by which mRNA directs the production of a _________ Modified image from What is Life? A Guide to Biology, Third Edition © 2015 W.H. Freeman and Company Transcription and Translation Transcription 1. In transcription, a section of DNA unwinds and nucleotides on DNA form base pairs with nucleotides of messenger RNA, creating an strand of mRNA. 2. This segment of mRNA then leaves the cell nucleus, headed for a ribosome in the cell’s cytoplasm, where translation takes place. 1 2 Translation 3. mRNA chain attaches to a ribosome, then transfer RNA molecules transport amino acids to ribosome. A chain of amino acids is linked together in the order specified by the mRNA sequence. 4. When the amino acid chain is finished and folded up, a protein is formed. 3 4 Transcription RNA polymerase 1. _________________ unwinds a region of the DNA double helix. 2. RNA polymerase begins assembling RNA nucleotides on the DNA template. 3. The completed portion of the RNA transcript separates from the DNA. 4. After the RNA transcript is released from the DNA, and the DNA is rewound into its original form. Transcription is completed. RNA Polymerase RNA polymerase: enzyme that unzips double stranded DNA and synthesizes the formation of mRNA RNA nucleotides RNA polymerase Direction of transcription Newly made RNA Template strand of DNA Protein Synthesis Codon = ____________ sequence of mRNA that codes for a specific amino acid Codons and Amino Acids 20 amino acids in humans 9 essential Multiple codons code for same amino acid Ex: Codons UCU, UCC, UCA, and UCG all code for Serine Start codon: AUG Amino acid: Methionine Stop codons: UAA, UAG, and UGA Modified image from What is Life? A Guide to Biology, Third Edition © 2015 W.H. Freeman and Company Check your Understanding DNA sequence mRNA sequence Amino acid sequence TA C A AT G C G A C G T G C Check your Understanding Translation 4. A polypeptide chain is produced. Translation Ribosomal RNA (rRNA): organelle that translates mRNA sequence by binding with tRNA with appropriate anti-codon and building polypeptide chain Large and small sub-units The “START sequence” of the mRNA (signified by sequence AUG) is recognized by a corresponding tRNA molecule and the two ribosomal subunits. Large subunit Small subunit Three binding sites A site: site where ribosome binds with tRNA P site: site where amino acids are ________ together E site: site where tRNA prepares for release from ribosome Modified image from What is Life? A Guide to Biology, Third Edition © 2015 W.H. Freeman and Company Transfer RNA Transfer RNA (tRNA): Binds with amino acids, then transfers amino acids to ribosome for polypeptide formation Anti-codon = set of three ribonucleotides complementary to a specific mRNA codon sequence Same as _____________ except for uracil/thymine replacement Image modified from What is Life? A Guide to Biology, Third Edition © 2015 W.H. Freeman and Company Review of RNA Types Translation: Steps 1 and 2 Step 1: mRNA strand binds with small subunit of ribosome. mRNA codon AUG signifies start point of polypeptide sequence. tRNA with Methionine (MET) amino acid (UAC anti-codon) attaches to codon. Step 2: Large ribosomal subunit connects with small subunit. Second tRNA arrives with amino acid and binds with A site within the ribosome Large subunit Translation: Steps 3 and 4 Step 3: Amino acids bond together and start to form the polypeptide chain. tRNA molecules shift to the right leaving A site vacant for next tRNA molecule Step 4: tRNA molecule within E site is released from the ribosome and a new tRNA molecule carrying an amino acid moves into the A site Termination Release factor binds to stop codon (UAA, UAG, UGA), and initiate hydrolysis the bond to the polypeptide Polypeptide is released Ribosome subunits separate Mass Production of Proteins A single mRNA can be translated ____________ by multiple ribosomes Check your Understanding 1. True or False: DNA replication and RNA synthesis both use the same polymerase to copy the DNA 2. True or False: Transcription takes places in the nucleus, while translation occurs in the cytoplasm 3. True or False: Transfer RNA (tRNA) carries the copied DNA out of the nucleus where it binds with a ribosome 4. True or False: Different codons can code for the same amino acid Check your Understanding 4. Which of the following contains the anti-codon? a. Ribosomal RNA b. Transfer RNA c. DNA d. Messenger RNA Check your Understanding 5. At which of the following binding sites are the amino acids linked together? a. A-site b. E-site c. P-site d. T-site Check your Understanding 6. Which of the following codons would cause the production of the polypeptide to be terminated? a. AUG b. UGA c. UAG d. UAA e. All of the above are stop codons Check your Understanding 7.Which of the following polypeptides would be made from the sequence below? DNA sequence: T A C A A G C G C T G A a. Tyr-Lys-Arg-stop b. Met-Phe-Gly-Gly c. Tyr-Phe-Ala-Gly d. Met-Phe-Ala-Thr e. Ine-Edt-Ost-Udy Check your Understanding 8. A DNA strand is reads A G T T C A . What are the corresponding anti-codons associated with it? a. A G T T C A b. U C A A G U c. A G U U C A d. U G A A C U Mutations Mutation: a _________ change in the base sequence of DNA May result in production of a nonfunctional protein Completely ________ Starting DNA Incorrect base pairing Mutation Point mutation Types of Mutations Point mutation: occur when one base pair in the DNA is substituted for another, or when one is inserted or deleted. Normal gene mRNA Protein Met Lys Phe Gly Ala Lys Phe Ser Ala Base substitution Point mutation Met Base deletion Frame shift mutation Met Missing Lys Leu Ala His Different Types of Point Mutations Name Definition Example Consequence Original DNA sequence Original polypeptide Silent Missense Change in nucleotide that does not change amino acid specified by codon Change in nucleotide that changes amino acid specified by codon Change in genetic code but not the resulting protein Change in primary structure of protein Nonsense Change in nucleotide that results in early stop codon Premature termination, incomplete polypeptide Frameshift Addition or deletion of nucleotide Reading frame is shifted, massive missense Sickle Cell Anemia Sickle cell anemia: genetic disorder caused by a single point mutation that results in a change in the structure of hemoglobin Gene Regulation Gene regulation: control of gene _____________ (protein production) Self regulating Proteins that regulate gene expression are transcribed and translated Three forms of gene regulation Transcription factors Promoters and enhancers 2. Micro-RNAs 3. Alternative splicing 1. Gene Regulation: Transcription Factors Transcription factors: proteins that either ________ ________ the binding of RNA polymerase to the template DNA Enhancer proteins can increase rate of transcription Gene Regulation: Micro-RNAs Micro-RNAs _______ with mRNA Slicer enzyme destroys mRNA Others inhibit translation by locking up mRNA before it comes in contact with a ribosome 1. A length of micro-RNA is transcribed from DNA. 2. Following processing, the micro-RNA binds to a protein complex in the cytoplasm. 3. A messenger RNA (mRNA) arrives at the micro-RNAprotein complex. 4. mRNA binds to the microRNA through base pairing. 5. The mRNA is cut in two by the Slicer enzyme within the protein complex. Gene Regulation: Alternative Splicing Introns: ___________ sequences ≈ 90% of gene length Exons: sequence expressed as proteins Alternative slicing: a process of editing a single primary transcript in different ways to yield multiple messenger RNAs Made possible by introns Gene Regulation: Alternative Splicing ATG TG G T TA AT T G C T TG C T TC C G G A C T TA A Complementary DNA Strand TA C A C C A AT TA A C G A A C G A A G G C C TG A AT T DNA Template for RNA Synthesis Transcription Unprocessed RNA Remove introns (bold bases) and splice the exons together before RNA leaves nucleus in the nucleus Processed RNA in the cytoplasm Translation Amino acid sequence Bioengineering Bioengineering or Biotechnology = using technology to control biological processes as a means of meeting societal needs ____________ organism = an organism whose genome has incorporated one or more genes from another species Genetically Modified Organism (GMO) + = Overview of Bioengineering Process 1. Desired section of DNA is chopped using ____________ enzymes 2. DNA fragments are amplified using polymerase chain reaction (PCR) 3. DNA fragments are inserted into __________ of bacteria or viruses 4. Bacteria or viruses containing desired DNA replicate and grow 5. Bacteria or viruses with desired DNA fragment are identified Chopping of DNA Restriction enzymes = enzymes from bacteria that are used to cut DNA into desired fragments Inserting the DNA __________ = extra chromosomal rings of bacterial DNA Replicate independently of chromosomes Move into bacteria cells and synthesize proteins Multiplying the Desired DNA Bacteria or viruses containing desired DNA fragment multiply very quickly increasing Abundance of desired protein is made by bacterial colony Genetically Modified Organisms (GMOs) Genetic engineering: the manipulation of a species genome in ways that do not normally occur in nature __________ selection: intentional reproduction of individuals in a population with a desired trait Creating Genetically Modified Organisms Corn plant destroyed by butterfly larvae (caterpillars). Bacterial gene coding for Bt crystals, which are poisonous to the caterpillars, is inserted directly into the corn plant’s DNA. Bt crystals—toxic to caterpillars—are now produced by the corn plant itself and present in all cells, reducing the amount of pesticides the farmer must use. Creating Genetically Modified Organisms Genes from multiple organisms can be combined to create a new variety Golden rice: rice containing gene for production of beta-carotene 25 times more vitamin A! Use of Genetically Modified Organisms (GMOs) GMO vs non GMO GMO Controversy Organisms we want to kill may become ___________ Organisms we don’t want to kill may be killed inadvertently GMOs are not tested or regulated adequately Loss of _________________ among crops Escape of GMO animals may impact natural populations Check your Understanding 1. True or False: All mutations result in an altered protein 2. True or False: All genes are constantly being transcribed and translated 3. True or False: A majority of the corn, soybeans, and cotton grown in the U.S has been genetically modified Check your Understanding 4. Which of the following is not one of the forms of gene regulation? a. Micro-RNA b. Transcription factors c. Mutations d. Alternative splicing Check your Understanding 5. Which of the following mutations are the result of an addition or deletion of a nucleotide? a. Silent mutation b. Missense mutation c. Nonsense mutation d. Frameshift mutation