Basic concepts of Genes, DNA & RNA

Course: BSc Biotech Sem-ISubject : Fundamentals of Biotechnology

Unit - 2

1

2

BREAKTHROUGH DISCOVERY

• In 1953, James Watson and Francis Crick discovered the double helical structure of the DNA molecule

1.

3

The discovery was announced on February 28, 1953

2.

4

The First Paper appeared in Nature on April 25, 1953

5

3.

6

Watson and Crick - Awarded Nobel Prize in 1962

4.

5.

7

DNA

A purine always links with a pyrimidine base to maintain the structure of DNA.Adenine ( A ) binds to Thymine ( T ), with two hydrogen bonds between them.Guanine ( G ) binds to Cytosine ( C ), with three hydrogen bonds between them.

6

87

9

Chemical Structure Of DNA

8

10

9

11

Nucleoside & Nucleotide, the fundamental building block of DNA

glycosidic bond

phosphoester bond

10

12

11

13

12

14

Ribose

13

15

DNA & RNA - Nucleotide Bases

16

Nucleotide

adenine

deoxyribose

PO4

17

Nucleotides14.

Chargaff's rule

• Chargaff's rules state that DNA from any cell of all organisms should have a 1:1 ratio (base Pair Rule) of pyrimidine and purine bases and, more specifically, that the amount of guanine is equal to cytosine and the amount of adenine is equal to thymine.

• They were discovered by Austrian chemist Erwin Chargaff

18

Relative proportions (%) of bases in DNA

Organism %A %G %C %T A/T G/C %GC %AT

φX174 24 23.3 21.5 31.2 0.77 1.08 44.8 55.2

Maize 26.8 22.8 23.2 27.2 0.99 0.98 46.1 54

Octopus 33.2 17.6 17.6 31.6 1.05 1 35.2 64.8

Chicken 28 22 21.6 28.4 0.99 1.02 43.7 56.4

Rat 28.6 21.4 20.5 28.4 1.01 1 42.9 57

Human 29.3 20.7 20 30 0.98 1.04 40.7 59.3

Grasshopper

29.3 20.5 20.7 29.3 1 0.99 41.2 58.6

Sea Urchin 32.8 17.7 17.3 32.1 1.02 1.02 35 64.9

Wheat 27.3 22.7 22.8 27.1 1.01 1 45.5 54.4

Yeast 31.3 18.7 17.1 32.9 0.95 1.09 35.8 64.4

E. coli 24.7 26 25.7 23.6 1.05 1.01 51.7 48.3

19

20

CHEMICAL AND PHYSICAL PROPERTIES OF DNA

• ABSORPTION• The bases in DNA absorb ultraviolet

light at the wavelength of 260 nm• This absorption can be monitored using

a spectrophotometer• This is one method used to figure the

concentration of DNA in solution• The more DNA present, the higher the

absorption

21

• DENSITY• Density can be measured by CsCl-

density ultracentrifugation•  can be used to estimate G+C content• GC base pairs are more dense than AT

base pairs• Density studies show the existence of

satellite DNA

22

• DENATURATION• DNA is considered denatured when the

double stranded DNA molecule is converted into two single stranded molecules

• As thermal energy increases, the frequency of hydrogen bonds breaking between the molecules increases

• G-C base pairs are held together by three hydrogen bonds (A-Ts by two) and it therefore takes more energy (higher temperatures) to separate molecules with high GC contents

23

• Hydrophobicity of solvent• Hydrophobic substances will allow the

bases in DNA to dissolve into the solvent

• Whereas hydrophilic substances will keep the bases of DNA stacked upon one another in the orientation that most favors hydrogen bonding between DNA strands

24

• pH• Acidic pH cause breakage of

phosphodiester bonds between nucleotides and breakage of the N-glycosidic bond between the sugar and purine bases

• Alkali - Above pH 11.3, all hydrogen bonds are disrupted and the DNA is totally denatured

• Salts will stabilize the DNA double helix

25

• Electrophoresis• DNA has a negative charge that is proportional

to its size• This is due to the negatively charged

phosphates in the sugar-phosphate backbone• If DNA is placed in an electrical field it will

migrate towards the positive electrode (the cathode)

• smaller pieces will migrate faster than larger pieces

• Larger pieces have trouble squeezing through the gel matrix and are hence retarded while smaller pieces migrate easier

26

• Type of gels• Agarose is used to separate fairly large

DNA molecules– 5 million to a few thousands base pairs

• Polyacrylamide is used to separate small pieces of DNA– 2 to several hundred base pairs

• The size of DNA is estimated by comparing its migration through the gel to DNA molecules of known size

27

A, B and Z DNA• A form – favored by

RNA• B form – Standard

DNA double helix under physiological conditions

• Z form – laboratory anomaly, – Left Handed– Requires Alt. GC– High Salt/ Charge

neutralization 15.

2816.

2917

30

RNA

Three major classes of RNA:

Difference between RNA & DNARNA DNA

RNA nucleotides contain ribose sugar

DNA contains deoxyribose

RNA has the base uracil DNA has the base thymine

presence of a hydroxyl group at the 2' position of the ribose sugar.

Lacks of a hydroxyl group at the 2' position of the ribose sugar.

RNA is usually single-stranded

DNA is usually double-stranded

31

mRNA

• Transcripts of structural genes.

• Encode all the information

necessary for the synthesis of a

polypeptide of protein.

• Intermediate carrier of genetic

information; deliver genetic

information to the cytoplasm.

18

32

mRNA to Amino Acid Dictionary

19

33

20

34

tRNA

RNA molecules 70- 100

nucleotide long.

The secondary structure of the

tRNA resembles a D loop,

anticodon loop, and T loop and

the acceptor stem.

Carry correct amino acids to

their position along the mRNA

template to be added to the

growing polypeptide chain.

21

35

rRNA

• The central component of

the ribosome.

• Ribosome; factory for

protein synthesis;

composed of ribosomal RNA

and ribosomal proteins.

• rRNA provides a

mechanism for decoding

mRNA into amino acids.

• rRNA interact with the tRNAs during translation by providing peptidyl transferase activity. 22

36

RNA Helices are short, bulges, loops

23

37

Organization of DNA in eukaryotes

24

38

• Chromosome – – double stranded DNA

molecule packaged by histone & scaffold proteins

DNA double helix

nucleosome

30nm fiber

condensed chromosome

25

39

Genome

Gene• Is the basic units of

inheritance; it is a segment within a very long strand of DNA with specific instruction for the production of one specific protein.

• Genes located on chromosome on it's place or locus.

Genome and Gene

• Totality of genetic information of an organism.• Encoded in the DNA (for some viruses, RNA).

26

40

Image refrences

1.http://www.bbc.co.uk/history/historic_figures/crick_and_watson.shtml

2. http://m.harunyahya.com/tr/works/4159/Once-Upon-A-Time-There-Was-Darwinism/chapter/4732/Once-there-was-no-knowledge-of-biological-information

3. http://arabian-explorers.com/james-watson-francis-crick

4.https://www.bcps.org/offices/science/secondary/forensic/DNA.pdf

5.http://www.timerime.com/en/event/209438/Nobel+Prize+for+Literature/

6. http://pixgood.com/dna-model-labeled-structure.html

41

Image refrences7.http://www.yourarticlelibrary.com/dna/

deoxyribonucleic-acid-dna-model-chemical-composition-and-transformation-experiments/22676/

8.http://ircamera.as.arizona.edu/NatSci102/NatSci/text/nucleicacid.html

9. http://quizlet.com/11685525/chapter-8-slides-flash-cards/

10.http://biology200.gsu.edu/houghton%2704/4564%20%2714/lecture6.html

11. http://www.slideshare.net/oreonixha/dna-11921134

12. http://www.bvgh.org/Current-Programs/Neglected-Disease-Product-Pipelines/Global-Health-Primer/Targets/cid/ViewDetails/ItemID/18.aspx

42

Image refrences13. http://smarttechno.net/tag/purine-bases14. http://en.wikipedia.org/wiki/Nucleotide15.http://biomedical.youngdong.ac.kr/sgkim/

files/2013/06/POB_5e_ch08-SGKim.pdf16.

https://lh3.ggpht.com/b7SCiLAza-c_D_wbjzOP86F4AnXyKu_wrviHfbF3GFqCWJXzuqHQvIG7P51jXvJcNJnNfQ=s144

17.https://lh5.ggpht.com/uE2KYozDJ302Z1ScuRJwtmqjjthxvILPxYnYyxog8q75VSahpvcMgtztJl8qGrXKhgsugA=s136

18. https://lh4.ggpht.com/1cWohlnifUAe4-WcqCCzkt3_GjfAmlbW1Tc7D0rcQXm9j5qW6Ns_xRv11VPwLy1aOsRkaQ=s170

43

Image refrences19.https://lh4.ggpht.com/

u0_RqejooRi2Vz7xOVJ7UkYuzUZquHOQnm4hSncWyNEOF0nAvJcM4WQlkYPuWaf2JiFN=s101

20. https://lh3.ggpht.com/ma2QHNXrMmoFjD-RtySlsVEPZjKqVEh7dv2oytsVIXO570HJZUXABdFoleQeALL8wy1fbg=s85

21.https://lh4.ggpht.com/rp027AgQNGhK99zk9t5qoLYRJgES8MDj0tnom_OYqQbMbTepfE1UttFktW5mSDBKuzUQ=s85

22.https://lh4.ggpht.com/iFsbqDCWZz_IqdhnffJBMcOFPvy60NzdBMG9V8SF4hIzzPYpgOJj4uhhnOct-vCGCB1k=s85

23. http://people.csail.mit.edu/donglai/24.

http://www.jeansforgenesday.org/disorders/basicterms

25. http://carolguze.com/text/442-1-humangenome.shtml

26. http://science.howstuffworks.com/life/cellular-microscopic/cell7.htm

44

References

• Biochemistry by S C Rastogi• Genes XI by Krebs, Goldstein and

Kilpatrick • Biotechnology by H K Das • Biotechnology by John E. Smith (5th Ed.) • http://www.genomicglossaries.com/• http://biology-pages.info