nucleic acids

NUCLEIC ACIDS
- polymers that consist of monomers called nucleotides
• polymer: large molecule made up of chains of small molecular subunits called
monomers
• dehydration synthesis: formation of polymers
- nucleic acids store and transmit genetic information
- generally found inside the nucleus
- primary structure of proteins is encoded in the sequence of DNA that make up the
genes
CHROMOSOMES VS. CHROMATIN
- when the cell is dividing, the DNA is in the form of chromosomes
- when it isn’t, it is in the form of chromatin
TYPES OF NUCLEIC ACIDS
- deoxyribose acid (DNA) and ribonucleic acid (RNA)
- made up of nucleotides
NUCLEOTIDES
- consists of a pentose sugar, a nitrogenous base, and a phosphate group
- DNA and RNA differ in the type of sugar as well as in nitrogenous bases
- to indicate position, carbons are numbered
- ribose is a pentose sugar because it has 5 carbons
NITROGENOUS BASES
- single ring: pyrimidine (Cytosine, Thymine, Uracil)
- double ring: purines (Adenine and Guanine)
NAMING BASES
- example -> base: Adenine
- base + sugar = adenosine (nucleoside)
- base + sugar + phosphate = adenosine monophosphate (nucleotide)

DISCOVERY OF NUCLEOTIDES
1. FRIEDRICH MIESCHER
- A Swiss scientist who discovered a unique substance found in the nuclei of atoms
which he called nuclein
2. ALBRECHT KOSSEL
- German biochemist who discovered the basic components of nuclein
- deoxyribose, phosphate and the bases AGCT
- discovered a kind of nuclein that had ribose and Uracil instead of Thymine
- Nuclein was later renamed to nucleic acid and the two kinds were identified
3. PHOEBUS LEVENE
- Russian chemist who came up with the correct arrangement of the phosphate, sugar
and bases in a nucleotide
- discovered that the sugar and phosphates form the backbone of nucleic acid strand
via phosphodiester bonds
4. JAMES WATSON AND FRANCIS CRICK
- came up with the structure of DNA as a double helix by connecting the observations
and discoveries of other scientists
5. IRWIN CHARGAFF
- discovered that adenine = thymine and guanine = cytosine (roughly)
- demonstrated that there is complementary base pairing
6. ROSALIND FRANKLIN
- how many strands were present in DNA and how were they connected
- earned a doctoral degree in Physical Chemistry at Cambridge University and studied
in Paris at the Laboratoire Central where she learned X-ray diffraction techniques
- returned to England as a research associate in John Randall’s Lab @ London where
she crossed paths with Maurice Wilkins
- she came close to solving DNA structure using her X-ray crystallographic
photographs

CHARACTERISTICS OF DNA
- it is a double helix made up of two strands
- there are four types of nucleotides present (A,G,T,C)
- the sugar and the phosphate form the backbone (via phosphodiester bonds)
- the bases lie between the backbone and strands are held together by H-bonds
between the bases (A-T: 2 H bonds, G-C: 3 H bonds)
BASE PAIRING
- a particular purine can only pair up with a specific kind of pyrimidine in which
hydrogen bonds can be formed
- DNA strand has directionality -> one end is diff. from the other end (5’ and 3’ ends)
- the 2 strands are anti-parallel -> they run in opposite directions, this results to
complementarity of strands which is important to DNA replication
DIRECTIONALITY OF DNA
- the orientation of the carbon atoms of the pentose sugar
- there are 5’ and 3’ ends of the DNA (the last trailing carbon atoms)
ANTI-PARALLEL STRANDS
- nucleotides in DNA backbone are bonded from phosphate to sugar between 3’ and 5’
carbons
- DNA molecule has “direction” and complementary strand runs in opposite direction
 DNA replication
DNA REPLICATION
- produces an exact copy of DNA and utilizes several enzymes
- prior to cell division, DNA replication occurs in order to ensure that resulting cells will
each have an exact copy of the original DNA
- makes use of base pairing and a template
NUCLEOTIDES AS LANGUAGE
- the language of genes that we will see translated to the language of amino acids in
proteins
INFORMATION TRANSFER
- gene: the sequence of nucleotides within a portion of DNA that codes for a peptide or
a functional RNA
- sum of all genes = genome

COPYING DNA
- base pairing allows each strand to serve as a template for a new strand
- new strand is 1/2 parent template and 1/2 new DNA
- replication begins in a replication bubble
FIRST STEP: UNWIND
- using the helicase enzyme to unwind part of the DNA helix
- stabilized by single-stranded binding proteins (SSB)
SECOND STEP: BUILD
- build daughter DNA strand
- add new complementary bases using DNA polymerase III
- can only add nucleotides to 3’ end of a growing DNA strand
- needs a “starter” nucleotide to bond to
- stand only grows 5’ to 3’ end
ENERGY OF REPLICATION
- nucleotides arrive as nucleosides
- DNA bases with P-P-P (energy for bonding)
- DNA bases arrive with their own energy source for bonding
- bonded by enzyme DNA polymerase III

LEADING AND LAGGING STRANDS

- limit of DNA polymerase III is that it can only build onto 3’ end of existing DNA strand
- lagging strand (okazaki fragments) -> joined by ligase (“spot welder” enzyme)
- leading strand has continuous synthesis
RNA PRIMER
- built by primase
- serves as starter sequence for DNA polymerase III
DNA POLYMERASE 1
- removes sections of RNA primer and replaced with DNA nucleotides
- DNA polymerase I still can only build onto 3’ end of existing DNA strand
CHROMOSOME EROSION
- all DNA polymerases can only add to 3’ end of an existing DNA strand
- there are loss of bases at the 5’ ends in every replication
- chromosomes get shorter with each replication -> limit to ~50 cell divisions
TELOMERES
- repeating, non-coding sequences at the end of chromosomes (protective cap)
TELOMERASE
- enzyme that extends telomeres
- can add DNA bases at the 5’ end
- different level of activity in different cells (high in stem cells and cancer cells)

DNA POLYMERASES
1. DNA POLYMERASE III
- by Roger Kornberg (2006)
- 1000 bases per second -> leads to a lot of typos
- main DNA builder
2. DNA POLYMERASE I
- by Arthur Kornberg (1959)
- 20 bases per second
- editing, repairing and primer removal
- proofreads and corrects typos (repair mismatched bases and remove abnormal
bases which repair damage throughout life)
- reduces error rate from 1 in 10,000 to 1 in 100 million bases
3. FAST AND ACCURATE
- takes less than 1 hour to copy 5 million base pairs in its single chromosome
• divide to form 2 identical daughter cells
- human cell copies its 6 million bases and divide into daughter cells in only a few
hours
• remarkably accurate
• only ~1 error per 100 million bases and ~30 errors per cell cycle
 RNA transcription
TRANSCRIPTION
- uses DNA template to build RNA
- from DNA nucleic acid language to RNA nucleic acid language
- DNA -> proteins -> cells -> bodies
CENTRAL DOGMA
- flow of genetic information in a cell
- DNA -> mRNA -> protein -> trait
METABOLISM
- taught us about genes through inheritance of metabolic diseases
- suggested that genes coded for enzymes
- each disease (phenotype) is caused by non-functional gene product
• lack of enzyme, tay sachs, PKU (phenylketonuria), albinism

GEORGE BEADLE AND EDWARD TATUM

- one gene:one enzyme hypothesis
- genes act by regulating definite chemical events

RNA
- ribose sugar, single stranded, Uracil base instead of Thymine
- lots of RNAs (mRNA, tRNA, rRNA, siRNA)
TRANSCRIPTION
- making mRNA 5’ to 3’
- transcribed DNA strand is the template strand
- untranscribed DNA strand is the non-coding strand (same sequence as RNA)
- synthesis of complementary RNA strand @ transcription bubble
- enzyme used is RNA polymerase
STAGES OF TRANSCRIPTION
- all the stages of transcription occur inside the nucleus
1. INITIATION
- enzymes unzip the DNA double helix, exposing the template strand
- RNA polymerase attaches to the promoter as well as proteins called transcription
factors
2. ELONGATION
- RNA polymerase moves along the DNA template strand, adding RNA nucleotides that
are complementary to the exposed bases on the DNA template strand
3. TERMINATION
- a terminator sequence in the DNA signals the RNA polymerase to separate from the
DNA template and release the newly produced RNA
RNA POLYMERASES
- each has a specific promoter sequence that it recognizes
1. RNA POLYMERASE 1
- only transcribes rRNA genes; makes ribosomes
2. RNA POLYMERASE 2
- transcribes genes into mRNA
3. RNA POLYMERASE 3
- only transcribes tRNA genes

PROMOTER REGION
- binding site before beginning of gene
- TATA box binding site
• binding site for RNA polymerase and transcription factors
INITIATION COMPLEX
- transcription factors bind to promoter region
• suite of proteins which bind to DNA
• turn on or turn off transcription
- trigger the binding of RNA polymerase to DNA
MATCHING BASES OF DNA AND RNA
- match RNA bases to DNA bases on one of the DNA strands, making an RNA copy
- RNA polymerase 2 joins RNA nucleotides together into a strand of RNA
TERMINATOR SEQUENCE
- where terminator occurs; RNA polymerase and mRNA detach from the DNA, which
returns to its double helix form
- product is an RNA copy of a gene

EUKARYOTIC GENES
- eukaryotic genes are not continuous, they have junk
- exons -> the real genes; expressed/coding DNA
- introns -> the junk; noncoding inbetween sequence
MRNA SPLICING
- post-transcriptional processing which edits out introns
- to make mature mRNA transcript; single base added or lost throws off reading frame
- eukaryotic mRNA needs work after transcription
- primary transcript = pre-mRNA
snRNPs
- small nuclear RNA, proteins
SPLICEOSOME
- several snRNPs that recognize the splice site sequence which cuts and pastes genes
POST-TRANSCRIPTIONAL PROCESSING
- need to protect mRNA on its trip from nucleus to cytoplasm because enzymes in the
cytoplasm attack mRNA
- protect the ends by adding 5’ GTP cap and poly-A tail (Adenine, longer tail, mRNA
lasts longer, more protein is produced)

MRNA CODING
- mRNA codes for proteins in triplets (codons)
CRICK
- determined the 3-letter (triplet) codon system
NIRENBERG (47) AND KHORANA (17)
- determined mRNA-amino acid match
- added fabricated mRNA to test tube of ribosomes, tRNA and amino acids
- created artificial UUUUU…mRNA and found that UUU coded for phenylalanine
THE CODE
- the strongest support for a common origin for all life
- code is redundant -> several codons for each amino acid; 3rd base “wobble”
- start codon: AUG (methionine)
- stop codons: UGA, UAA, UAG
TRANSFER RNA STRUCTURE (tRNA)
- “clover leaf” structure; anticodon on clover leaf end
- amino acid attached on 3’ end
AMINOACYLE TRNA SYNTHETASE
- enzyme which bonds amino acid to tRNA
- bond requires energy
• ATP -> AMP
• bond is unstable, it can release amino acid at ribosome easily

RIBOSOMES
- facilitate coupling of tRNA anticodon to mRNA codon
- ribosomal RNA (rRNA) and proteins
- two subunits: large and small
A SITE (aminoacyl-tRNA site)
- holds tRNA carrying next amino acid to be added to chain
P SITE (peptidyl-tRNA site)
- holds tRNA carrying growing polypeptide chain
E SITE (exit site)
- empty tRNA leaves ribosome from exit site
BUILDING A POLYPEPTIDE
1. INITIATION
- brings together mRNA, ribosome subunits, initiator tRNA
2. ELONGATION
- adding amino acids based on codon sequence
3. TERMINATION
- end codon
 DNA technology
HUMAN GENOME
- 3.2 million bases
DNA TECHNOLOGY
- change DNA to add new traits or create a modified or new organism
GENETICALLY MODIFIED ORGANISMS (GMOs)
- also called transgenic organisms; genes are transferred from one organism to
another

ARTIFICIAL SELECTION
- breeders choose which organism to mate to produce offspring with desired traits
- cannot control what genes are passed on
1. SELECTIVE BREEDING
- organisms with the desired characteristics are mated to produce offspring with
those desired traits
- desired genes are passed on to the next generation (ex. dog breeds)
2. HYBRIDIZATION
- two individuals with unlike characteristics are crossed to produce best of both
- Luther Burbank created a disease resistant potato called Burbank potato -> he
crossed a disease resistant plant with one that had large food producing capacity
- Ex. Grape + Apple = Grapple
3. INBREEDING
- breeding of organisms that are genetically similar to maintain desired traits
- dog breeds are kept pure this way, keeps each breed unique from others
- risk: since both have same genes, there is a higher chance to get recessive genetic
disorder (ex. blindness, joint deformities)

STEM CELLS
- undifferentiated animal cells that can give rise to specialized cell types
- embryonic stem cells are totipotent -> come from very early embryos and can give
rise to all cell types in the body
- adult stem cells are pluripotent -> come from adult tissues and can give rise to many
but not all cell types in the body
CLONING
- technique in which an identical copy of an organism can be made
- identical twins are naturally created clones (splitting of an embryo into two)
- clone: group of cells or organisms that are genetically identical as a result of asexual
reproduction; they will have the same exact DNA as their “parent”
- common in nature among single-celled organisms such as bacteria and many
protists -> plants are very easy to clone
CLONING IN ANIMALS
- single cell is removed from a parent organism
- using culture media and chemicals, a new individual will be grown from that cell
- one cell has all DNA needed and each cell in the body has the same DNA but cells vary
because different genes are turned on in each cell
DOLLY
- the first cloned mammal
- somatic cell was removed from the breast of Dolly’s mother and nucleus was
extracted and placed into an egg cell of another sheep
- egg cell was induced to become an embryo and then implanted into uterus of egg
donor sheep -> dolly had exact same DNA as her mother and had no father
HOW TO CLONE HUMAN
- egg cell is removed from female and nucleus of the egg is removed
- somatic cell is removed from other person and nucleus is removed
- nucleus of somatic cell is placed into the egg so that egg no longer needs to be
fertilized since it will have 46 chromosomes
- egg is charged with electricity to start mitosis and then put into surrogate mother
GENE SPLICING
- direct manipulation of DNA
- desirable genes can be extracted from one organism and transferred to another

BACTERIA
- one-celled prokaryotes; dominant form of life on Earth
- reproduce by mitosis (binary fission)
- rapid growth (generation every 20 minutes; 100 million colony overnight)
BACTERIAL GENOME
- single circular chromosome; haploid
- naked DNA with no histone proteins
- around 4 million base pairs and 4300 genes, 1/1000 DNA in eukaryote
BACTERIA ARE OPPORTUNISTS
- pick up naked foreign DNA wherever it may be hanging out
- have surface transport proteins that are specialized for uptake of naked DNA
- import bits of chromosomes from other bacteria
- incorporate the DNA bits into their own chromosome to express new genes,
transform and recombine
PLASMIDS
- small supplemental circles of DNA that can be imported from environment
- 5000 to 20,000 base pairs that self-replicate
- carry extra genes (2-30) for antibiotic resistance
- can be exchanged between bacteria (rapid evolution)
- a way to get genes into bacteria easily
• insert new gene into plasmid
• insert plasmid into bacteria = vector
• bacteria now expresses new gene to make new protein
RESTRICTION ENZYMES
- restriction endonucleases by Werner Arber, Daniel Nathans, Hamilton Smith
- evolved in bacteria to cut up foreign DNA
• restrict the action of the attacking organism
• protection against viruses and other bacteria
• bacteria protect their own DNA by methylation and not using the base sequences
recognized by the enzymes in their own DNA
- cut DNA at specific sequences (restriction site)
- symmetrical “palindrome”; produces protruding ends (sticky ends) that will bind to
any complementary DNA
- many different enzymes names after organism they are found in
STICKY ENDS
- cut other DNA with same enzymes and can glue DNA together
MIXING GENES
- gene produces protein in different organisms or different individuals

TRANSGENIC PLANTS
- Recombinant TI plasmids (tumor inducing) combine with plant chromosomes when
plant cells are infected by genetically modified bacteria
- plant develops tumors which contain infected cells carrying desired gene
- these infected cells would be grown (via cloning) into new plants
- Ex. protect crops from insects (BT corn), extend growing season (fishberries),
improve quality of food (golden rice)
TRANSGENIC ANIMALS
- genetically modified viruses carrying desired genes can infect early embryonic cells
- when cells divide by mitosis, they pass along recombinant DNA to all cells

DNA SEQUENCING
- helps deduce protein sequences and determine evolutionary relationships
- makes use of the process of electrophoresis
POLYMERASE CHAIN REACTION (PCR)
- rapidly produces millions of copies of a DNA sequence in a test tube (to analyze)
- target DNA strands are separated by heat and polymerases synthesize new
complementary strands to match them
- number of molecules of DNA doubles with every round of PCR
- can detect genetic differences between individuals by amplifying DNA
DNA PROFILING
- examines short tandem repeats (STRs) which are short DNA sequences from the
noncoding regions of DNA -> comparing them reveals if they are a match
- the STRs are passed along from parent to child same way as genes
DNA PROBES
- can test mutations from any gene in an embryo, child or adult
PREIMPLANTATION GENETIC DIAGNOSIS (PGD)
- to reduce the odds of having a child with a genetic disease
- a man’s sperm fertilizes several eggs (in vitro fertilization)
- extract DNA from one cell of each embryo, amplify it using PCR and use DNA probes
to search for harmful alleles
- DNA can be screened to identify genetic diseases
GENE THERAPY
- may someday provide new treatment options for genetic diseases
- goal is to supplement a faulty gene with a normal, healthy version
CRISPR-CAS9
- uses guide RNA to find the gene they want to edit
- guide RNA is coupled to an enzyme that cuts out DNA and replaces it with different
sequence
- quickly and easily modify one specific gene without affecting other part of genome