Lab 1: Bioinformatics

Amino acids

organic molecules that are protein building blocks; they contain an amino group, a carboxyl group, and a unique side chain.

DNA

Deoxyribonucleic acid; double-stranded nucleic acid that carries genetic information, has nucleotide bases (ATCG) & a sugar/phosphate backbone, stores/transmits hereditary information— codes, decodes, regulates, and expresses genes.

RNA

Ribonucleic acid; single-stranded nucleic acid that helps with gene expression (e.g. mRNA, tRNA, rRNA) and protein synthesis, carrying messages from DNA to ribosomes for translation.

mRNA

Messenger RNA; type of RNA that carries genetic info from DNA to ribosome; then translated into protein.

Codon

triple-nucleotide-base-sequence in mRNA that specifies an amino acid or signals end of translation

Contig

continouous sequence of DNA assembled from overlapping fragments during genome sequencing; used to create a consensus sequence for the genome.

Base pair

complementary nitrogenous base pair in DNA or RNA (DNA: A-T, C-G)

Transcription

RNA is synthesized from DNA template, making mRNA that carries genetic instructions for protein synthesis.

Translation

ribosomes synthesis proteins based on codon sequences in mRNA

Steps to protein synthesis

transcription, mRNA processing, translation, & post-translational modifications.

Allele

variant form of gene that arises due to mutations and contributes to genetic diversity in a population, resulting in different traits.

Genomic DNA Library

DNA fragment collection with an organism’s entire genome, stored in vectors for research & sequencing

Complementary DNA (cDNA)

DNA copy synthesized from mRNA template using enzyme “reverse transcriptase” — represents only expressed genes since introns are removed

Intron

non-coding sequence of RNA w/in a gene that’s removed during mRNA processing before translation

Exon

coding sequence in gene that stays in mRNA after splicing & is translated into protein

Genetic code

rule set that defines how nucleotide sequences in mRNA are translated into amino acid sequences in proteins (codons)

genome

complete set of organism’s genetic material (genes + introns)

Protein

biomolecule made of amino acids linked by peptise bonds; have structural, enzymatic, & regulatory functions in cells

Shotgun Sequencing

genome sequencing where DNA is randomly fragmented, sequenced, & reassembled computationally

Nonsense Mutation

where codon is changed into stop codon — premature termination of protein synthesis

Polymorphism

DNA variation in DNA sequence that occurs in 1% of population & contributes to geentic diversity w/out necessarily causing disease

Single nucleotide polymorphisms (SNPs)

single base-pair variation in DNA that occurs & used as geentic marker for traits/diseases

Coding strand

(sense strand) DNA strand with same sequence as mRNA (except thymine is replaced by uracil in RNA) and serves as reference during transcription. RNA polymerase reads emplate strand to make mRNA sequence identical to coding strand.

Template strand

antisense strand - DNA that is reference for RNA synthesis; RNA polymerase reads this strand in 3’ to 5’ direction to make mRNA in 5’ to 3’ direction (coomplementary to template strand)

UTR (Untranslated Region)

noncoding sections of mRNA that regulate gene expression w/out being translated into proteins

• 5’ UTR: before start codon; regulates translaion initiaition

• 3’ UTR: after stop codon; influences stabilitiy, localization, & translation efficiency

Nucleic Acids

biomolecules that store & transmit genetic information (phosphate group, 5-C sugar, nitrogenous base (ATCG)) - Ex: DNA & RNA

Phosphodiester bond

3’ to 5’ - covalent bonds btw nucleotides in the backbone of DNA & RNA

Bioinformatics

Study of collecting, sorting, analyzing DNA & protein sequence information using computers & statistical techniques

Bioinformatics components

1. database creation

2. development of algorithms & statistics

3. use of tools for analysis/interpretation of biological data

EX: DNA/protein sequences, protein structures, image processing, gene expression profiles, & biochemical pathway

Fast methods for sequencing databases, RNA, & proteins

Dideozy sequencing (Sanger method), cycle sequencing, mass spectrometry

Protein sequencing obtained?

X-ray crytallography, NMR Spectroscopy

Global expression analysis?

DNA microarrays, RNA-seq

Types of sequence database

Genbank (NCBI)

Types of protein structure databases

PDB, SWISS-PROT

Organism-specific resources

EcoGene, OMIM

Specialized sequence databases

rRNA, tRNA, promoter sequences

Methods of bioinformatics

DNA sequence assembly, sequence alignment & gene finding, reading frame translators (6-frame translation), protein sequence comparisons (pair-wise & multiple aignments).

What are bioinformatics used in?

• drug design & discovery

• genome assembly & transcriptomic profiles

• protein structure alighnment & prediction

• modeling of evolution

Sequence assembly

public domain software (assemblers (assemble DNA sequnce data into contigs. Software merges short fragments of DNA to reconstrcut the original seuence.

ORF finder

graphical analysis tool that finds all open reading frames in a sequence (start codon - AUG - to Stop codon - UAA, UAG, UGA)

BLAST

• basic local alignment search tool

• computer program that searches for gene sequence similarities - can be used to identify homologus genes in different organisms

Clustal Omega

compares many DNA and amino acid sequences (multiple sequence alignments) & aligns them to highlight their similarities

• can give important information of proteins & evolutionary relationships (phylogenetic trees)

Nucleotide BLAST

• includes gene name, species & methodology related to how it was sequenced

• Max score: calculation includes matches & mismatches, along w/ other data (higher max score = better alignment)

• E-value: indicates statistical significance of the data (lower e-value = more significance)

Protein BLAST

• Domains: distinct functional and/or structural units in protein; often responsible for specific functions/interactions, contributing to overall protein role

• Alignment: shows which amino acids are the same & which are different btw the query and hit

Difference btw cDNA & Genomic DNA clones

cDNA clones are sued for studying gene expression & making recombinant proteins - genomic DNA clones are used for studying gene regulation, mutation, & genomic structure

How many alpha chains and how many beta chains are present in a normal adult hemoglobin tetramer?

2 of each

What disease is caused by reduced amounts of detectable beta globin?

Beta-plus-thalassemia

What disease is caused by absence of beta chain?

Beta-zero-thalassemia

On which chromosome is the human HBB gene located?

11th - p-arm

How to read phylogenetic tree

• nodes/branch points indicate divergence from last common ancentsor - sharing immediate branch point means higher similarity (vs ones that branched multiple points back)

• branch lengths are proportional to amount of change