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Copyright © 2004 by Limsoon Wong A Biology Review Body • Our body consists of a number of organs • Each organ is composed of a number of tissues • Each tissue is composed of cells of the same type Cell • Performs two types of function – Chemical reactions necessary to maintain our life – Pass info for maintaining life to next generation • In particular – Protein performs chemical reactions – DNA stores & passes info – RNA is intermediate between DNA & proteins Protein • A protein sequence composed from an alphabet of 20 amino acids – Length is usually 20 to 5000 amino acids – Average around 350 amino acids • Folds into 3D shape, forming the building blocks & performing most of the chemical reactions within a cell Classification of Amino Acids • Amino acids can be classified into 4 types. • Positively charged (basic) – Arginine (Arg, R) – Histidine (His, H) – Lysine (Lys, K) • Negatively charged (acidic) – Aspartic acid (Asp, D) – Glutamic acid (Glu, E) Classification of Amino Acids • Polar (overall uncharged, • Nonpolar (overall but uneven charge uncharged and uniform distribution. can form charge distribution. cant hydrogen bonds with form hydrogen bonds water. they are called with water. they are hydrophilic) called hydrophobic) – – – – – – – Asparagine (Asn, N) Cysteine (Cys, C) Glutamine (Gln, Q) Glycine (Gly, G) Serine (Ser, S) Threonine (Thr, T) Tyrosine (Tyr, Y) – – – – – – – – Alanine (Ala, A) Isoleucine (Ile, I) Leucine (Leu, L) Methionine (Met, M) Phenylalanine (Phe, F) Proline (Pro, P) Tryptophan (Trp, W) Valine (Val, V) Genetic Code • Each amino acid is composed of three nucleotides • Start codon: ATG (code for M) • Stop codon: TAA, TAG, TGA Copyright © 2004 by Limsoon Wong DNA • DNA stores instruction needed by the cell to perform daily life function • Consists of two strands interwoven together and form a double helix • Each strand is a chain of some small molecules called nucleotides Francis Crick shows James Watson the model of DNA in their room number 103 of the Austin Wing at the Cavendish Laboratories, Cambridge Copyright © 2004 Limsoon Wong Classification of Nucleotides • 5 different nucleotides: adenine(A), cytosine(C), guanine(G), thymine(T), & uracil(U) • A, G are purines. They have a 2-ring structure • C, T, U are pyrimidines. They have a 1-ring structure • DNA only uses A, C, G, & T A C Copyright © 2004 by Limsoon Wong G T U Watson-Crick rules • Complementary bases: – A with T (two hydrogen-bonds) – C with G (three hydrogen-bonds) C A T 10Å Copyright © 004 by Limsoon Wong G 10Å Double Stranded DNA • DNA is double stranded in a cell. The two strands are anti-parallel. One strand is reverse complement of the other • The double strands are interwoven to form a double helix Copyright © 2004 by Limsoon Wong Locations of DNAs in a Cell? • Two types of organisms – Prokaryotes (single-celled organisms with no nuclei. e.g., bacteria) – Eukaryotes (organisms with single or multiple cells. their cells have nuclei. e.g., plant & animal) • In Prokaryotes, DNA swims within the cell • In Eukaryotes, DNA locates within the nucleus Chromosome • DNA is usually tightly wound around histone proteins and forms a chromosome • The total info stored in all chromosomes constitutes a genome • In most multi-cell organisms, every cell contains the same complete set of chromosomes – May have some small differences due to mutation • Human genome has 3G base pairs, organized in 23 pairs of chromosomes Gene • A gene is a sequence of DNA that encodes a protein or an RNA molecule • About 30,000 – 35,000 (protein-coding) genes in human genome • For gene that encodes protein – In Prokaryotic genome, one gene corresponds to one protein – In Eukaryotic genome, one gene can corresponds to more than one protein because of the process “alternative splicing” Complexity of Organism vs. Genome Size • Human Genome: 3G base pairs • Amoeba dubia (a single cell organism): 600G base pairs Genome size has no relationship with the complexity of the organism Number of Genes vs. Genome Size • Prokaryotic genome (e.g., E. coli) – Number of base pairs: 5M – Number of genes: 4k – Average length of a gene: 1000 bp • Eukaryotic genome (e.g., human) – Number of base pairs: 3G – Estimated number of genes: 30k – 35k – Estimated average length of a gene: 1000-2000 bp • ~ 90% of E. coli genome are of coding regions. • < 3% of human genome is believed to be coding regions Genome size has no relationship with the number of genes! RNA vs DNA • RNA is single stranded • Nucleotides of RNA are similar to that of DNA, except that have an extra OH at position 2’ – Due to this extra OH, it can form more hydrogen bonds than DNA – So RNA can form complex 3D structure • RNA use the base U instead of T – U is chemically similar to T – In particular, U is also complementary to A Central Dogma • Gene expression consists of two steps – Transcription DNA mRNA – Translation mRNA Protein Copyright © 2004 by Limsoon Wong Transcription • Synthesize mRNA from one strand of DNA – An enzyme RNA polymerase temporarily separates doublestranded DNA – It begins transcription at transcription start site (ATG) – A A, CC, GG, & TU – Once RNA polymerase reaches transcription stop site, transcription stops (TGA, TAG, and TAA) • Additional “steps” for Eukaryotes – Transcription produces pre-mRNA that contains both introns & exons – 5’ cap & poly-A tail are added to pre-mRNA – RNA splicing removes introns & mRNA is made – mRNA are transported out of nucleus Translation • Synthesize protein from mRNA • Each amino acid is encoded by consecutive seq of 3 nucleotides, called a codon • The decoding table from codon to amino acid is called genetic code • 43=64 diff codons Codons are not 1-to-1 corr to 20 amino acids • All organisms use the same decoding table • Recall that amino acids can be classified into 4 groups. A single-base change in a codon is usually not sufficient to cause a codon to code for an amino acid in different group Ribosome • Translation is handled by a molecular complex, ribosome, which consists of both proteins & ribosomal RNA (rRNA) • Ribosome reads mRNA & the translation starts at a start codon (the translation start site) • With help of tRNA, each codon is translated to an amino acid • Translation stops once ribosome reads a stop codon (the translation stop site) Introns and exons • Eukaryotic genes contain introns & exons – Introns are seq that are ultimately spliced out of mRNA – Introns normally satisfy GT-AG rule, viz. begin w/ GT & end w/ AG – Each gene can have many introns & each intron can have thousands bases • Introns can be very long • An extreme example is a gene associated with cystic fibrosis in human: – Length of 24 introns ~1Mb – Length of exons ~1kb Typical Eukaryotic Gene Structure Image credit: Xu • Unlike eukaryotic genes, a prokaryotic gene typically consists of only one contiguous coding region Copyright © 2004 by Limsoon Wong Reading Frame • Each DNA segment has six possible reading frames Forward strand: ATGGCTTACGCTTGA Reading frame #1 Reading frame #2 Reading frame #3 ATG GCT TAC GCT TGC TGG CTT ACG CTT GA. GGC TTA CGC TTG A.. Reverse strand: TCAAGCGTAAGCCAT Reading frame #4 Reading frame #5 Reading frame #6 TCA AGC GTA AGC CAT CAA GCG TAA GCC AT. AAG CGT AAG CCA T.. Copyright © 2004 by Limsoon Wong Open Reading Frame (ORF) • ORF is a segment of DNA with two in-frame stop codons at the two ends and no in-frame stop codon in the middle stop stop ORF • Each ORF has a fixed reading frame Coding Region • Each coding region (exon or whole gene) has a fixed translation frame • A coding region always sits inside an ORF of same reading frame • All exons of a gene are on the same strand • Neighboring exons of a gene could have different reading frames Frame Consistency • Neighbouring exons of a gene should be frame-consistent ATG GCT TGG GCT TTA A -------------- GT TTC CCG GAG AT ------ T GGG exon 1 exon 2 exon 3 exon1[i, j] in frame A and exon2[m, n] in frame B are consistent if B = (m - j - 1 + A) mod 3 What is Gene Finding? • Find all coding regions from a stretch of DNA sequence, and construct gene structures from the identified exons • Can be decomposed into – Find coding potential of a region in a frame – Find boundaries between coding & non-coding regions Search-by-Homology Example: Gene Finding Using BLAST • High seq similarity typically implies homologous genes Search for genes in yeast seq using BLAST Extract Feature for gene identification candidate gene Image credit: Xu BLAST search Genbank or nr Copyright © 2004 by Limsoon Wong sequence alignments with known genes, alignment p-values • Searching all ORFs against known genes in nr db helps identify an initial set of (possibly incomplete) genes sequence BLAST hits Image credit: Xu known nongenes % 0 known genes coding potential gene length distribution • A (yeast) gene starts w/ ATG and ends w/ a stop codon, in same reading frame of ORF • Have “strong” coding potentials, measured by, preference models, Markov chain model, ... • Have “strong” translation start signal, measured by weight matrix model, ... • Have distributions wrt length, G+C composition, ... • Have special seq signals in flanking regions, ...