Bioinformatics Flashcards
How to use plasmids?
1-Design: Select a suitable plasmid vector
2- Construction: Introduce desired DNA sequences
3- Verification: Confirm the successful construction of the recombinant plasmid.
4- Transformation: Transfer the recombinant plasmid into the target organism.
5- Expression: Cultivate transformed cells t
Selectable marker (Marker gene)
Gene conferring antibiotic resistance trait
Multiple cloning site (MCS)
Region with multiple restriction enzyme recognition sites for inserting foreign DNA.
Reporter Gene
A reporter gene is a gene whose activity serves as a visible or measurable indicator of the activity of other genes.
PCR
What: Technique to amplify DNA fragments.
Why: Replicate DNA for various analyses.
How: Cyclically heat, cool, and extend DNA using primers and DNA polymerase.
Primer design
What: Designing primers that flank a target DNA region for use in PCR.
Why: To amplify specific DNA sequences for various applications such as DNA sequencing and cloning.
How: Utilizing bioinformatics tools to select primer sequences with appropriate length, melting temperature, and specificity to the target DNA region.
Restriction Enzymes -Sticky/Blunt
What: Enzymes that cut DNA at specific recognition sequences.
Why: Used in genetic engineering to manipulate DNA.
How: Recognize specific DNA sequences and cleave them at specific sites.
Ligation
What: Joining DNA fragments together.
Why: Used in molecular biology to create recombinant DNA molecules.
How: DNA ligase enzyme catalyzes the formation of phosphodiester bonds between DNA fragments.
Types of primers for different purposes.
-Normal PCR. [(18-30 bp)]
-Parenthood type PCR. [(Short <15bp)]
-RE- Primers. [(Adding the RE sequence to the primer’s end)]
-Overlapping primers. [(Complement the other primer back)]
Design primers need some specs
1-Types of primers for different purposes.
2- Thermodynamics.
3- GC content.
4- AT ending
5- Length of the primers.
6- Primers dimers and self dimers
Overlapping PCR
Overlapping PCR involves designing primers with overlapping ends to amplify DNA fragments, which can then be joined together to create a fused DNA sequence.
(PCR Done with fragments that have Tails complement to each other)
Extension PCR
PCR done using special primers with Tail
Purification PCR
PCR to amplify the full constructed fragment
Genomics in bioinformatics
involves using computational tools and techniques to analyze, interpret, and manage large-scale genomic data efficiently.
Genomics is a branch of molecular biology that focuses on the:
Structure,
Function,
Evolution,
Mapping,
And Editing of Genomes
Analyzes entire genome, providing comprehensive genetic information.
Whole Genome Sequencing (WGS)
It makes use of high-throughput techniques, such as as WGS, to sequence DNA fragments rapidly and in parallel.
Next-Generation Sequencing (NGS):
Gene structure [Open reading Frame ORF] CDS:
Represents the portion of a gene that potentially encodes a protein product.
Gene orientation
Refers to the directionality of a gene on a DNA strand, either forward (+) or reverse (-) relative to a reference point.
Gene mining
Involves the identification and extraction of genes or genetic information from genomic data.
Annotation
The process of identifying and labeling genes, regulatory sequences, and other functional elements within a genome.
Comparative Genomics
Analyzes genome similarities/differences to understand evolution, functions, and variations.
Phylogenetics
Reconstructs evolutionary relationships using molecular data to build evolutionary trees.
for Gene location (Locus), use:
NCBI, KEGG, UNIPROT
For Gene orientation use:
SNAPGENE, Negative strain
for Relationship between organisms
MEGA software
Biosynthetic gene clusters (BGCs)
Involves identifying the sets of genes responsible for the synthesis of specialized metabolites, such as antibiotics or natural products, within the genomic sequence of microorganisms.
Transcriptomics
is the study of all the RNA molecules present in a cell, tissue, or organism at a given time.
-It provides insights into which genes are active and how they are regulated, offering valuable information about cellular processes and functions
Next Generation Sequencing (NGS) steps:
1- Extraction
2-library preparation
3-sequencing
4-Analysis
Basics of Transcriptomics Analyses?
1-Gene Expression Analysis
2-Sequence Alignment
3-Functional Annotation
4-Differential Gene Expression Analysis
5-Visualization of Transcriptomic Data
Studying the levels of RNA molecules produced from different genes in a biological sample.
Gene Expression Analysis
The process of arranging DNA or RNA sequences to identify similarities and differences to identify mutations
Sequence Alignment
Assigning biological functions to genes based on their sequence characteristics or experimental evidence to roles of genes in various cellular processes.
Functional Annotation
Analysis that compares gene expression levels between sample groups to identify genes that are significantly upregulated or downregulated.
Differential Gene Expression (DGE) Analysis
Visualization tools allow researchers to explore and interpret transcriptomic data in a more intuitive manner patterns.
Visualization of Transcriptomic Data
Gene Ontology
it’s a standardized system for categorizing genes based on their biological functions, molecular roles, and cellular locations
a biomedical and genomic information hub through databases, tools, and resources for researchers, healthcare professionals, and the public.
NCBI
Tool for comparing DNA and protein sequences.
NCBI BLAST
Kyoto Encyclopedia of Genes and Genomes, a database for biological pathways, diseases, drugs, and genomes.
KEGG
Deep learning-based protein structure prediction software developed by DeepMind, known for its accuracy in determining protein structures from amino acid sequences
AlphaFold Database
Search Tool for the Retrieval of Interacting Genes/Proteins, a database that compiles known and predicted protein-protein interactions.
String Database
Arrangement of sequences to identify similarities and differences, often used in bioinformatics to compare DNA, RNA, or protein sequences.
Offline Alignments
Molecular Evolutionary Genetics Analysis, software used for conducting molecular evolutionary analysis, phylogenetic tree construction, and Alignment.
MEGA
Molecular visualization software used for viewing and analyzing molecular structures in 3D.
PyMol
UGENE
a versatile software tool, is a go-to choice for bioinformatics tasks
