10: Biotechnology and Microscopy

microscopy

allows us to see objects invisible to the naked eye (cellular level)

optical microscope

directs visible light through optical lenses

electron microscope

directs beam of electrons for greater magnification

compound microscopy

visible light focused on a thin sample, producing 2D image; can see many cells, tissues, and organisms

compound microscopy

disadvantage of ___: samples thicker than one cell layer require staining, which kills specimen

florescence microscopy

florescent probes used to tag structures; can see where specific molecules, like proteins, are located in the cell

florescence microscopy

advantage of ___: florescent markers do not kill sample, allowing live viewing

compound and florescence

types of optical microscopy

Scanning Electron Microscopy (SEM —> S for Surface)

electron beams shot across surface of sample, producing high-resolution 3D images of surface; visualizes the exterior of very small objects, like ribosomes

scanning electron microscopy

disadvantage of ___: sample must be dehydrated and coated before viewing, which kills specimen

transmission electron microscopy (TEM)

electron beams shot through a thin section of sample, producing high-resolution 2D images; visualizes internal structures of tissues, cells, and organelles

transmission electron microscopy

disadvantage of ___: samples must be dehydrated, fixed, and sliced, which keeps specimen

transmission electron microscopy

What type of microscopy uses the highest magnification microscope?

cell fractionation

process by which cells are lysed and the contents are separated based on density and size

homogenization

cell membrane is disrupted and the mixture of contents (homogenate) is collected

differential centrifugation

successive spinning at low, medium, and high speeds to create pellets of decreasing density

nuclei, mitochondria/chloroplast, ribosomes/viruses

Relative size and density of cellular components (largest to smallest)

vertical gene transfer

gene transfer from one generation to the next (sexual/asexual reproduction, mitosis)

horizontal gene transfer

gene transfer between different, unrelated organisms; can occur naturally or via biotechnology

conjugation, transduction, transformation

3 types of horizontal gene transfer

conjugation

DNA transferred directly from one cell to another vie a pilus (bacterial appendage used to exchange cytoplasm and small molecules)

transduction

DNA transferred from cell to cell by a virus

transformation

DNA taken up by cell from environment; to transform a cell, it must first be made competent to promote spontaneous DNA uptake via heat-shock or electroporation (processes that increase membrane permeability)

recombinant DNA

DNA which contains segments from different sources; DNA segments can be exchanged via horizontal gene transfer or using artificial technology

restriction enzymes

recombinant DNA technology uses ___ to cut DNA at sequence-specific, palindromic “recognition sites”

restriction mapping

creating a map of all enzyme recognition sites present in a piece of DNA

restriction fragment length polymorphisms (RFLPs)

different individuals will have differing nucleotide sequences, containing recognition sites at different locations in their DNA. digestion with restriction enzymes produces unique DNA fragment patterns and lengths for each individual

DNA fingerprinting

use RFLPs to match an individual’s DNA to a sample

gel electrophoresis

separates DNA, RNA, or proteins based on size and charge

negative; positive

In gel electrophoresis, DNA/RNA travel from __ to __ electrode.

further

In gel electrophoresis, shorter/smaller molecules move ___ than longer molecules.

ssDNA (radioactively-labeled single-stranded DNA) probes

What is added to locate specific sequences in gel electrophoresis?

SDS

Proteins are treated with ___ before being loaded into the gel, which denatures and adds negative charge to proteins.