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BIO205-CH9-rDNAtech
BIO205 - CH9 - Recombinant DNA Technology - Tortora - Rio Salado - AZ
| Question | Answer |
|---|---|
| How can microorganisms & plants be used as "factories" to produce unnatural chemicals/products? | By inserting genes into cells by recombinant DNA tech (rDNA) - genetic engineering |
| Another name for recombinant DNA (rDNA) technology. | genetic engineering |
| vector | A plasmid or virus used in genetic engineering to insert genes into a cell. |
| DNA vectors are often called __. | gene-cloning vectors, or cloning vectors. |
| Site-directed mutagenesis. | Techniques used to modify a gene in a specific location to produce the desired polypeptide. |
| Site-directed mutagenesis can be used to __. | make a specific change in a gene. |
| restriction enzymes | Special class of DNA - cutting enzymes that exist in many bacteria. |
| Restriction enzymes protect bacteria from __. | a hydrolyzing phage DNA - protected because cell methylates (add methyl groups). |
| methylates | Using restriction enzymes to add a methyl group to cytosines in DNA. |
| What role do restriction enzymes play? | Recognizes & digests (cuts) only are particular sequence of nucleotide bases in DNA. |
| What is useful about sticky ends left by staggered cuts? | They "stick" to complementary stretches of single-stranded DNA by base-pairing. Same enzymes used on 2 different sources of DNA will splice (recombine) in vitro. |
| Order of formation of rDNA molecule. | (1) restriction enzyme cuts 2 DNA at its particular site, (2) sticky ends formed, (3) sticky ends bond w/hydrogen bonds, (4) DNA ligase covalently bonds backbones of DNA pieces to produce rDNA molecules. |
| Why are restriction enzymes used to make recombinant DNA? | Because they produce staggered cuts that create "sticky ends" that can bond w/other DNA strands cut by same enzymes. |
| Most important property of a vector. | self-replication |
| What does an ori (origin of replication) allow a plasmid to do? | be self-replicating |
| Vectors serve as __. | vehicles for the replication of desired DNA sequences. |
| Which size of vector is easier to manipulate? | Smaller vectors, because larger DNA molecules are more fragile. |
| Most important property of a vector. | self-replication |
| What does an ori (origin of replication) allow a plasmid to do? | be self replicating |
| Vectors serve as __. | vehicles for the replication of desired DNA sequences. |
| Which size of vector is easier to manipulate? | Smaller vectors, because larger DNA molecules are more fragile. |
| What is special about shuttle vectors? | They are plasmids that are capable of existing in several different species & can be used to move cloned DNA sequences among speces. |
| Viral DNA | Vector that can usually accept much larger pieces of foreign DNA than plasmids can. |
| Name some viruses that are used as viral vectors. | Retro viruses, adenoviruses, & herpes viruses. |
| Polymerase chain reaction (PCR) | Technique by which small samples of DNA can be quickly amplified (increased quantities). |
| PCR can amplify __. | small samples of DNA |
| primers | Short pieces of nucleic acid added to help start reactions - complementary to ends of target DNA & hybridize fragments. |
| After each cycle of synthesis, DNA is __ to convert all the new DNA into a single strand. | heated |
| What enzyme copies DNA in PCR? | DNA polymerase |
| DNA polymerase for PCR comes from __. | Thermus aquaticus - a thermophilic bacterium - allows for heating by a thermalcycler. |
| PCR __ be used to amplify an entire genome. | cannot |
| In genetic engineering, a plasmid is inserted into a cell by __. | transformation |
| transformation | Procedure during which cells can take up DNA from environment - cells soaked in Ca chloride & made competent - heat added & then they take up DNA. |
| electroportation | Using electric current to create micro pores in membrane - cells then absorb DNA through pores. |
| protoplasts | Cell walls removed enzymatically to make bacteria vulnerable to experimentation. |
| Protoplast fusion | Protoplasts in a solution will fuse & create hybrid cell - can merge species like plant & alge. |
| Microinjection | inserting DNA into a cell w/glass micropipette |
| Foreign DNA will only survive if it __. | is present on self-replicating vector or is incorporated into a chromosome by recombination. |
| gene library | collection of clones containing different DNA fragments. |
| Genes are cloned into vectors using __. | restriction enzymes |
| The enzyme reverse transcriptase is used to synthesize __. | complementary DNA (cDNA) |
| cDNA is synthesized from __ template. | an mRNA |
| A DNA copy of mRNA is produced by __. | reverse transcriptase |
| cDNA | DNA made in vitro from an mRNA template. |
| The __ method is most common method of obtaining eukaryote genes. | cDNA |
| How does cDNA differ from synthetic DNA. | Synthetic DNA is created mechanically by assembling nucleotides as if letters of a keyboard, while cDNA contains exons from a mRNA template. |
| Blue-white screening for clones | If a bacterium received the original plasmid containing a piggy-backed gene, it will hydrolyze x-gal & produce a blue-colored compound. |
| Colony hybridization | ID of a colony containing a desired gene by using DNA probe complementary to that gene. |
| DNA probe | Short, labeled, single strand of DNA or RNA used to locate its complementary strand in a quantity of DNA. |
| DNA probes are labeled with? | A radioactive element of fluorescent dye. |
| In blue-white screening, white contains? | Foreign DNA |
| In blue-white screening, blue contains? | no foreign DNA |
| Why is E. coli used in genetic modifications. | They are grown easily & their genetics are known. |
| What is one disadvantage of using E. coli for genetic engineering. | They are gram-negative & produce endotoxins. |
| Why use yeast cells in genetic engineering? | They are eukaryotic & may carry plasmids. |
| gene silencing | Defense against viruses & transposons. |
| RNA interference (RNAi) | Target a particular gene - like a virus gene - and binds to mRNA & destroys it - silencing the expression of the gene. |
| One technique for genome sequencing is __. | random shotgun sequencing. |
| Random shotgun sequencing | Technique for determining the nucleotide sequence in an organism's genome. |
| bioinformatics | The science of understanding the function of genes through computer-assisted analysis. |
| proteomics | Science of determining all of the proteins expressed in a cell. |
| 2 uses of Southern blotting | Genetic screening & DNA fingerprinting. |
Created by:
Ladystorm
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