Eukaryotic cell: 20

micrometer in diameter.

Nucleus is 5μm in diameter.

23 pairs of DNA , the average

of which is 3×109bp, amount

to more than 1.7 meters of

DNA.

The structure of Eukaryotic Chromosomes

chromatin (染色质)

the interphase in cell divisions

(细胞分裂间期)

chromosome (染色体)The mitotic time

(细胞分裂期)

Nucleosomes Are the Fundamental

Organizational Units of Chromatin

Nucleosome: The binding of DNA by the

histones forms a structure unit of chromatin,

the structure unit is called nucleosome.

Nucleosome

Linker

region

Nucleosome

core

Histone H1

60bp of DNA

Octamer of histone (two

copies each of H2A 、H2B、H3、H4)

146bp of DNA

Octameric core

DN

A w

rap

ped

aro

un

d a

his

ton

e co

re

Histones Are Small, Basic Proteins

*The sizes of these histones vary somewhat from species to species.

The numbers given here are for bovine histones.

Nucleosomes. Regularly spaced nucleosomes consist

of histone octamer bound to DNA.

(a) Schematic illustration (b) electron micrograph.

(beads-on-a-string)

nucleosome

H1 histone

Histone octamer

nucleosome

Nucleosomes Are Packed into SuccessivelyHigher Order Structures

Produce 40-60-fold compaction of DNA

7 fold compaction from DNA to nucleosome

螺线管

The 30 nm fiber, a higher-order organization of nucleosomes. (a) Schematic illustration of the probable structure of the solenoid fiber, showing nucleosome packing. (b) Electron micrograph.

Compaction of DNA in a eukaryotic chromosome

超螺线管螺线管核小体 染色质纤维

7 40-60 40

IV. Function of DNA:

*DNA is carrier of genetic information.

It can transfer genetic information by replication.

*The genes in DNA can be transcribed into RNA

that are involved in the synthesis of protein.

* By definition, gene is a functional unit of DNA.

A gene represents a genetic information that can

be transcribed into RNA ,which are involved in

the synthesis of protein.

Section Ⅲ

Structures and Functions of RNA

The chemical feature of RNA differs from that of DNA

RNAs are much smaller than DNA.

In RNA, the sugar moiety is ribose rather than

2’-deoxyribose of DNA.

Instead of thymine, RNA contains the uracil.

RNA exists as a single strand, whereas DNA exists as

a double-stranded helical molecule.

In RNA, guanine content ≠cytosine content; adenine

content ≠uracil content.

RNA can be hydrolyzed by alkali, whereas DNA not

Uracil forms base

pairs with adenin

Seco

ndary

str

uct

ure

of

RN

As

The Classification of RNA

Coding RNA----mRNA

Noncoding RNA

Constitutive noncoding RNA

tRNA, rRNA, snRNA,

snoRNA, scRNA

Regulatory noncoding RNA

lncRNA, miRNA, circRNA,

siRNA, piRNA

In the prokaryotic and eukaryotic organisms,

there are three main classes of RNA

mRNA (messenger RNA)

tRNA (transfer RNA)

rRNA (ribosomal RNA)

Each of them differs from the others by size,

function and general stability.

I. Structure and function of mRNA

Serve as templates of protein synthesis.

The most heterogeneous in size and

stability.

Unique characteristics (in eukaryote):

cap of 5’-terminal (7-mGpppNm)

tail of 3’-terminal(polyA)

Eukaryotic mRNA molecule

5’ UTR and 3’UTR: are related to the regulation

of translation

The 5’ cap of mRNA

7-Methylguanosine is

joined to the 5’ end of

almost all eukaryotic

mRNAs in an unusual

5’,5’-triphosphate

linkage. Methyl groups

(pink) are often found

at the 2’ position of the

first and second

nucleotides. m7GpppNm-

m7GpppNm-

Capping

Methylation

5’-5’ triphosphate linkage

加帽酶

function

Bind with cap-binding

complex of protein，(CBC)

Prevent from degradation

Participates in

exportation from the

nucleus

Participates in translation

initiation

hnRNA：heterogeneous nuclear RNA, precursor mRNA

Intron: the noncoding regions, can not express into

protein sequence.

Exon: the encoding regions is a part of the gene that,

can represent protein sequence.

hnRNA contains the encoding

region and noncoding region

hnRNAs are processed to generate mRNAs

hnRNA

Note: One triplet code

denote one amino acid.

The “message” in mRNA is carried in

groups of three nucleotides called triplet code:

2. 3'-Polyadenylation

A tail of A-nucleotides,

generally 100-200 long, is added

to the 3‘-end of eukaryotic or

most prokaryotic pre-mRNAs.

features:

not coded by template DNA;

*added in nucleus;

*retained in the mRNA;

*functions in mRNA stability

and translation activity.

Involved in transportation of mRNA

Helps stabilize the mRNA

The 5’ cap and 3’polyA can protect mRNA from

ribonucleases.

Involved in recognition and binding with ribosome in

initiation of translation

Functions of 5’cap ( 7-mGpppNm) and 3’ poly A tail

The cap and tail participate in binding of the mRNA to

the ribosome to initiate translation

The 3’ and 5’ ends of

eukaryotic mRNAs

are linked by a

complex of proteins

that includes several

initiation factors and

the poly(A) binding

protein (PAB). This

complex binds to the

40S ribosomal

subunit.

Serve as carrier（adapters) of aa in the

translation.

Vary in length from 73 to 93 nucleotides .

Contains more rare bases.

II . Structure and function of tRNA

There are at least 32 species of tRNA

molecules in every cell, each kind of

tRNA carry one kind of corresponding

amino acid, so at least one (and often

several) correspond to each amino acids.

The tRNA Molecules Serve as

Adapters for Translation

General cloverleaf secondary structure of tRNAs

The large dots on the

backbone represent

nucleotide residues; the blue

lines represent base pairs.

Characteristic and/or

invariant residues common

to all tRNAs are shaded in

pink. tRNAs vary in length

from 73 to 93 nucleotides.

Extra nucleotides occur in

the extra arm or in the D

arm.

dihydrouridine

D loop

T ψC loop

anticodon loop

4 major loops and 1 arm

D /DHU loop

TψC loop

Anticoden loop

amino acide loop

extra arm

amino acid arm of tRNA

bind a specific amino acid

anticodon arm recognize

the nucleotide sequence

encoding that amino acid

in an mRNA.

AUC→isoleucine

The Pairing Relationship of Codon and Anticodon

tertiary structure of tRNA deduced from x-ray

diffraction analysis.

The shape resembles a twisted L. (a) Schematic diagram with

the various arms shaded in different colors. (b) A space-filling

model, with the same color coding.The CCA sequence at the 3

end (orange) is the attachment point for the amino acid.

III. Structure and Function of rRNA

rRNAs are the components of ribosome.

A ribosome is a cytoplasmic nucleoprotein

structure that acts as the machinery for the

protein synthesis.

4 types

3 types

Diagrams of the

secondary structure of E.

coli 16S and 5S rRNAs.

The first (5’ end) and

final (3’ end)

ribonucleotide residues

of the 16S rRNA are

numbered

The predicted secondary structure of the Bacterial rRNAs

Non-coding RNAs

Consititute non-coding RNA

ribozyme

small nucleolar RNA snoRNA

small nuclear RNA snRNA

small cytoplasmic RNA scRNA

Regulatory non-coding RNA

small non-coding RNA (snRNA)

micro RNA miRNA

small interfering RNA siRNA

pivi interacting RNA piRNA

large non-coding RNA (lncRNA)

circular RNA (circRNA)

Section Ⅳ Chemical Features of

Nucleic Acid and Applications

Conformation: long linear molecule

Viscosity: highly viscous at pH 7.0 and

room temperature (25℃)

dsDNA 〉ssDNA

DNA 〉RNA

Acidity: highly acidity

Absorbance for UV light at 260nm

Ⅰ. General features of nucleic acid:

The spectra are shown as the variation in molar extinction coefficient with

wavelength. The molar extinction coefficients at 260 nm and pH 7.0 (ε260) are

listed in the table. The spectra of corresponding ribonucleotides and

deoxyribonucleotides, as well as the nucleosides, are essentially identical. For

mixtures of nucleotides, a wavelength of 260 nm (dashed vertical line) is used for

absorption measurements.

Absorption spectra of the common nucleotides

≥

≥

Definition: DNA denaturation means that a DNA has

lost its’ native conformation, and double strands DNA is

separated to single strand DNA by exposed to a

destabilizing factors such as heat, acid, alkali,urea , et al.

(when high temperature is used to denature DNA, the

DNA is said to be melted).

Ⅱ. Denaturation of DNA

Definition of Tm:

Tm is melting

temperature at which

half (50%) of DNA

molecules are

denatured.

Heat denaturation of DNA

The denaturation/melting curves of two DNA specimens.

The temperature at the midpoint of the transition (Tm) is

the melting point

Influence factors of Tm

1.it depends on pH and ionic strength

2. the size of DNA

3. base composition of the DNA:

the G/C content is higher,

the DNA melting point is higher

Tm as a characteristic for DNA denaturation

•Tm is proportional to (G+C)% of DNA molecules

Tm=69.3+0.41(G+C)%

Tm=4(G + C) + 2(A + T) (＜20bp)

•Tm also depends on DNA length

Relationship between Tm and GC content of DNA

DNA renaturation :

Removing the denaturation factors or in proper condition,

the denatured DNA (ssDNA) restore native structure

(dsDNA) and function. The renaturation of heat

denatured DNA by slowly cooling is called annealing.

Ⅲ. Renaturation

Reversible denaturation and annealing (renaturation)

of DNA

Hyperchromic effect and hypochromic effect:

Ⅳ Nucleic Acids from Different Species

Can Form Hybrids

Definition of Hybridization:

when heterogeneous single strand DNA or RNA are

put together, they will become to heteroduplex via the

base-pairing rules during renaturation if they are

complementary in parts (not complete). This is called

molecular hybridization.

Significance of hybridization:

. Hybridization can be used to detect similar DNA

sequences in two different species. This reflects a common

evolutionary heritage. The closer the evolutionary

relationship between two species, the more extensively

their DNAs will hybridize

Application of hybridization

• molecular biology :Detect or isolation a specific DNA sequence or RNA in the presence of many other sequences with specific probe

Probe: a short appropriate complementary DNA strand (usually labeled in some way) to hybridize with the target DNA sequence. can be isolated from a different or the same species, or it can be synthesized chemically in the laboratory.

• Forencis science: make possible the identification of an individual on the basis of a single hair left at the scene of a crime

• Medicine: the prediction of the onset of a disease decades before symptoms appear

SectionⅤ

Specific Nucleases Digest Nucleic Acids

deoxyribonucleases

ribonucleases

Endonucleases: restriction endonucleases

exonucleases5′exonucleases

3′exonucleases

(DNase)

(RNase)

Nucleases Differ in Their Specificity for

Different forms of Nucleic Acids