Biotechnology 11

What is DNA Technology?

Isolation, purification, analyzation, and manipulation of DNA.

What is genetic cloning?

The impact of gene by isolating and replicating it.

What are cloning vectors?

Examples include bacterial plasmids used to isolate, replicate, and study genes.

What is biotechnology?

Techniques applied to biological systems for modification.

What does genomics involve?

Characterization of whole genomes.

What is basic research in genetics?

Study of gene structure, function, and regulation of expression.

What is applied research in genetics?

Other applications such as gene therapy.

What is a plasmid?

Circular double stranded DNA that can replicate independently of chromosomal DNA.

What function do plasmids serve?

Vectors for inserting genes.

What is recombinant DNA?

DNA fragments from two or more different sources joined into a single molecule.

What does CRISPR stand for?

Clustered Regularly Interspaced Short Palindromic Repeats.

What is the role of the RNA guide in CRISPR?

It is complementary to the target gene.

What is Cas9 in CRISPR technology?

A DNA endonuclease that cleaves the target so a new gene can be inserted. It is a guide

What does Cas9 recognize in the genome?

The PAM (Proto-spacer Adjacent Motif) sequence.
- 3 nucleotides

What is gene cloning?

Gene cloning involves DNA cloning that specifically targets a gene.

What are genetically modified organisms (GMOs)?

GMOs have their genomes engineered to introduce or change a genetically controlled trait.

What do GMOs contain?

GMOs contain recombinant DNA, which consists of fragments from two or more different sources that are joined together.

What are the uses of cloned DNA in research?

Cloned DNA is used in basic research (studying gene structure and function) and applied research (medical, forensic, agricultural, or commercial applications).

What are cloning vectors?

Cloning vectors are DNA molecules into which a DNA fragment is inserted to form a recombinant DNA molecule for cloning purposes.

What are plasmid cloning vectors engineered with?

They are engineered with the ampR gene and the lacZ gene to locate bacteria that incorporate recombinant plasmids.

What does the ampR gene do?

The ampR gene encodes an enzyme that breaks down the antibiotic ampicillin.

What does the lacZ gene encode?

The lacZ gene encodes β-galactosidase, which hydrolyzes the sugar lactose.

How are DNA molecules transformed in cloning?

DNA molecules are transformed into ampicillin-sensitive, lacZ – E. coli, which are spread on plates containing ampicillin and the β Galactosidase synthetic substrate X-gal.

What is blue-white screening?

Bacteria that have been transformed with recombinant plasmids are identified by blue-white screening.

How does gene cloning begin with plasmids?

Gene cloning involves cutting the plasmid's DNA to insert the gene of interest.

What are sticky ends in DNA cloning?

Sticky ends are DNA fragments with single-stranded ends that can hydrogen bond with complementary sticky ends.

What role does ligation play in DNA cloning?

Ligation involves DNA ligase sealing the sugar-phosphate backbones of DNA strands together.

What is a restriction endonuclease?

Restriction endonucleases are bacterial enzymes that cut DNA at specific sites known as restriction sites.

What is a restriction site?

A restriction site is a sequence of approximately six to eight base pairs that a given restriction enzyme recognizes and binds to.

What happens if a restriction enzyme cannot find a restriction site?

If a restriction enzyme cannot find a specific restriction site on a bacterial plasmid, it will not initiate a cleaving event.

What are restriction fragments?

Restriction fragments are DNA fragments produced by restriction enzymes that have sticky ends.

What is the significance of sticky ends in DNA cloning?

Sticky ends enable the joining of two DNA molecules cut with the same restriction enzyme through hydrogen bonding.

Where are restriction endonucleases primarily produced?

Restriction endonucleases are primarily produced in prokaryotes, such as bacteria, and not in human cells.

How do bacteria use restriction endonucleases?

Bacteria use restriction endonucleases to combat foreign infectious agents, cutting foreign nucleic acids into fragments for breakdown.

What is the Polymerase Chain Reaction (PCR)?

PCR is a technique used to make many copies of a specific segment of DNA without cloning the sequence in a host organism.

What is the primary purpose of PCR?

The primary purposes of PCR are DNA amplification and the identification of diseases by isolating specific sequences to amplify.

How many cycles does a typical PCR run?

A typical PCR cycle runs 20-30 times.

What is a primer in PCR?

A primer is a specific sequence used for complementary pairing to isolate the target DNA sequence for amplification.

What occurs during the denaturation stage of PCR?

During denaturation, the DNA target sequence, Taq polymerase, nucleotides, and primers are heated to 95°C to denature the DNA into single strands.

What happens during the annealing stage of PCR?

In the annealing stage, the temperature is cooled to 55-65°C, allowing primers to bind to complementary sequences on the target DNA sequence.

What occurs during the extension stage of PCR?

During extension, the temperature is raised to 72°C, where Taq polymerase replicates the target sequence, producing two identical copies of DNA.

What is Taq DNA polymerase?

Taq DNA polymerase is a highly thermostable enzyme from the thermophilic bacterium Thermus aquaticus, ideal for PCR due to its ability to operate at high temperatures.

Why is Taq polymerase important for PCR?

Taq polymerase is important for PCR because it can withstand high temperatures, enabling the denaturation and amplification processes to occur efficiently.

What is PCR typically used for?

PCR is typically used for amplifying and quantifying nucleic acids in various applications, including medical diagnostics and research.

What are introns?

Introns are segments of DNA found within a gene that do not code for proteins; they are removed during mRNA processing.

What are exons?

Exons are the parts of the gene that contain the actual instructions for making the protein.

What happens to mRNA after processing in eukaryotes?

After processing, a double-stranded DNA copy can be made from mRNA via reverse transcription.

What is reverse transcriptase?

Reverse transcriptase is the enzyme that synthesizes a complementary DNA (cDNA) copy from an RNA template.

What is complementary DNA (cDNA)?

cDNA is a copy of DNA synthesized from an mRNA template, which can be cloned and expressed in bacterial cells.

What is the role of reverse transcription in molecular biology?

Reverse transcription allows for the creation of DNA from mRNA, facilitating stability and replication for further study.

How does reverse transcription differ from the central dogma?

Reverse transcription reverses the central dogma by going from RNA back to DNA instead of from DNA to RNA to protein.

What are the steps in reverse transcription?

1. Isolate mRNA from the cell.
   2. Use nucleotide primers complementary to the Poly-A tail of mRNA.
   3. Reverse transcriptase synthesizes DNA complementary to mRNA.
   4. The mRNA strand is degraded.
   5. DNA polymerase assists in creating cDNA.

What is electrophoresis in molecular biology?

Electrophoresis is a technique used to separate DNA molecules based on their size, with longer DNA molecules having more base pairs and greater mass.

What is bioinformatics?

Bioinformatics is an interdisciplinary field that combines computer science, mathematics, physics, and biology to analyze and interpret biological data, leading to advances in various areas like stem cell research.

What are stem cells?

Stem cells are undifferentiated cells that have the potential to become any specialized cell type in the body.

What is totipotent stem cell?

Totipotent stem cells can differentiate into all cell types, including extraembryonic tissues such as the placenta.

What is pluripotent stem cell?

Pluripotent stem cells can differentiate into any cell type within the embryo but not into extraembryonic tissues.

What are multipotent stem cells?

Multipotent stem cells have a restricted ability to produce only certain types of cells; they commonly include adult stem cells.

What is the role of adult stem cells?

Adult stem cells are multipotent and function to replace specialized cells in various tissues and organs.

What is germline gene therapy?

Germline gene therapy involves the experimental introduction of a gene into germline cells, but it is currently not allowed in humans.

What is somatic gene therapy?

Somatic gene therapy is a medical technique that treats genetic disorders by modifying cells from a person's body and returning them to the individual to correct genetic issues.

What are CRISPR loci and Cas genes?

CRISPR loci and Cas genes are segments in bacteria that store DNA from past viral infections, enabling the bacteria to recognize and combat the viruses upon future attacks.

How do bacteria use CRISPR arrays?

Bacteria capture fragments of viral DNA, insert them into their own DNA to create CRISPR arrays, allowing them to 'remember' previous infections and respond effectively.

What role does Cas9 play in CRISPR?

Cas9 is an enzyme used by bacteria to cut apart viral DNA, which disables the virus and protects the bacterium from infection.

What is genetic engineering used for in agriculture?

Genetic engineering is used to produce transgenic crops that have altered traits, such as enhanced resistance to herbicides or pests.

What are Round-up ready crops?

Round-up ready crops are genetically modified to be resistant to Round-up, a herbicide containing glyphosate that disrupts protein synthesis necessary for plant growth.

What is the shikimic acid pathway?

The shikimic acid pathway is an essential enzyme pathway in plants and some microorganisms for synthesizing specific amino acids vital for growth.

What is plant pharming?

Plant pharming is the genetic modification of plants and animals to produce substances that can be used as pharmaceuticals or therapeutics.

What are restriction fragment length polymorphisms (RFLPs)?

RFLPs are DNA fragments of varying lengths that are derived from the same region of the genome and are analyzed to detect genetic variations.

How are RFLPs typically analyzed?

RFLPs are typically analyzed using agarose gel electrophoresis, which separates the DNA fragments based on size.

What is a single-nucleotide polymorphism (SNP)?

An SNP is a single base-pair mutation in a gene, such as the β-globin gene associated with sickle-cell syndrome.

What are DNA fingerprints?

DNA fingerprints are unique combinations and variations of DNA sequences in humans, identified through techniques like RFLPs and SNPs, except in identical twins.

What is DNA fingerprinting?

DNA fingerprinting is a technique using PCR to analyze DNA variations across various loci in the genome, often involving 13 loci in noncoding regions as standards.

What is a short tandem repeat (STR) sequence?

An STR sequence, or microsatellite, is a short DNA sequence repeated in series, with each repeat being 2 to 6 base pairs in length.

What is gel electrophoresis used for?

Gel electrophoresis is used to separate DNA, RNA, or protein molecules in a gel subjected to an electric field, based on size, charge, or other properties.

What is agarose gel electrophoresis specifically used for?

Agarose gel electrophoresis is specifically used to separate DNA molecules based on their size.

What does a Southern blot detect?

A Southern blot is a technique used for DNA detection.

What does a Northern blot detect?

A Northern blot is a technique used for RNA detection.

What does a Western blot detect?

A Western blot is a technique used for protein detection.

What are transgenic organisms?

Transgenic organisms are those that have received genes from an external source, resulting in new traits.

What is a knockout mouse?

A knockout mouse is a genetically engineered mouse with embryonic stem cells that have had a transgene added to induce a nonfunctional state of a specific gene.

What is gene therapy?

Gene therapy is the introduction of a normal gene into specific cell lines to correct genetic disorders.

What is the Ti plasmid?

The Ti (tumor-inducing) plasmid is a plasmid from the bacterium Agrobacterium tumefaciens that serves as a vector for making transgenic plants.

What is the role of Bt toxin in agriculture?

Bt toxin provides insect resistance in genetically modified crops, helping protect them from pests.