Biotechnology: Principles & Processes Flashcards

Comprehensive vocabulary flashcards covering principles, tools, steps, methods, and key enzymes of biotechnology based on Class 12 Biology NCERT notes.

Biotechnology (EFB Definition)

Defined by the European Federation of Biotechnology as "the integration of natural science and organisms, cells, parts thereof, and molecular analogues for products and services."

Genetic Engineering

Techniques to alter the chemistry of genetic material ($DNA/RNA$) to introduce into host organisms, thus changing the phenotype of the host cell.

Bioprocess Engineering

Maintenance of sterile (microbial contamination-free) conditions to enable growth of desired microbes/eukaryotic cells in large quantities for commercial production.

Exonucleases

Enzymes that remove nucleotides from the ends of the $DNA$.

Endonucleases

Enzymes that cut at specific positions within the $DNA$ strand.

Palindromic Recognition Sequences

Specific DNA sequences where the sequence reads the same in the $5' \rightarrow 3'$ direction on both complementary strands, targeted for cutting by endonucleases.

Action of EcoRI Enzyme

Cuts double-stranded $DNA$ specifically between G & A in the sequence $5'\text{-GAATTC-}3'$ and $3'\text{-CTTAAG-}5'$, leaving sticky ends.

EcoRI Naming Convention

Named after Escherichia (E), coli (co), strain RY13 (R), and 1st enzyme isolated (I). First letter from genus (Capital), next two from species (lowercase), next from strain, and Roman numeral indicates order of isolation.

Origin of Replication (ori)

A specific sequence in a vector where replication starts and which controls the copy number of the linked $DNA$.

Selectable Marker

Genes encoding resistance to antibiotics (such as $amp^R$ and $tet^R$ in pBR322) used to identify and select transformants from non-transformants.

Insertional Inactivation

The inactivation of a marker or coding gene (e.g., $\beta\text{-galactosidase}$) due to insertion of recombinant $DNA$, resulting in white colonies for recombinants and blue colonies for non-recombinants in the presence of chromogenic substrate.

pBR322

A cloning vector plasmid ($4361\text{ bp}$) featuring $amp^R$ and $tet^R$ marker genes, restriction sites (PstI, PvuI, EcoRI, ClaI, HindIII, BamHI, SalI), $ori$, and $rop$.

Chemical Method (Transformation)

A transformation technique for bacteria (E. coli) involving treatment with a divalent cation ($Ca^{2+}$), followed by heat shock ($42^\text{o}\text{C}$) and placement back on ice.

Micro-injection

A transformation method for animal cells where recombinant $DNA$ is directly injected into the nucleus using a glass micropipette.

Biolistics / Gene Gun

A transformation method for plant cells where cells are bombarded with high-velocity microparticles of gold or tungsten coated with $DNA$.

Disarmed Pathogens Method

Gene delivery method using modified Agrobacterium tumefaciens for plants or Retroviruses for animals to infect cells safely.

Chilled Ethanol Precipitation

A step in genetic material isolation where purified $DNA$ precipitates out as fine threads upon the addition of chilled ethanol.

Gel Electrophoresis

A process where negatively charged $DNA$ fragments move toward the Anode ($+$) through an agarose matrix under an electric field, with smaller fragments moving faster and traveling farther.

Ethidium Bromide (EtBr)

A stain used to visualize $DNA$ bands under UV light as bright orange-colored bands following gel electrophoresis.

Elution

The process of cutting out and extracting $DNA$ bands from an agarose gel.

Polymerase Chain Reaction (PCR)

An in vitro technique used to amplify a specific gene segment into billions of copies through Denaturation, Annealing, and Extension.

Denaturation (~94°C)

The first step of PCR where high temperature breaks hydrogen bonds, separating double-stranded $DNA$ into two single strands.

Annealing (~54°C)

The second step of PCR where two sets of oligonucleotide primers bind to complementary regions at the $3'$ ends of single $DNA$ strands.

Extension (~72°C)

The third step of PCR where thermostable Taq Polymerase extends primers using $dNTPs$.

Taq Polymerase

A thermostable $DNA$ polymerase isolated from Thermus aquaticus that remains active at high temperatures during the extension step of PCR.

Simple Stirred-tank Bioreactor

A culture vessel equipped with an impeller for continuous mixing and air availability, along with foam controller, temperature, and $pH$ control systems.

Sparged Stirred-tank Bioreactor

A bioreactor in which sterile air bubbles are sparged (bubbled) through the reactor chamber, dramatically increasing the surface area for gas exchange.

Downstream Processing

The collective operations after the biosynthetic stage, including separation, purification, formulation with preservatives, quality control, and clinical trials.

DNA Ligase

An enzyme originating from Bacteriophage or E. coli that joins sticky/blunt ends by forming phosphodiester bonds.

Alkaline Phosphatase

An enzyme originating from calf intestine or bacteria that prevents self-ligation of a vector by removing the $5'$ phosphate group.

Lysozyme / Chitinase

Enzymes originating from egg white (Lysozyme) or fungi (Chitinase) used to break bacterial or fungal cell walls during $DNA$ extraction.