BASIC PRINCIPLES OF DNA a short discussion

Basic principles of DNA

1. Basic Principles of Human Genetics By Dr. Tapan Kr. Mondal (DCH, MD)

2. Mendel’s theory of inheritance Law of segregation Law of dominance Law of independent assortment

3. Basic concept of human genetics The genetic information of an individual is contained in 23 pairs of chromosomes. Every human cell contains 23 pairs of chromosomes. 1 pair is called sex chromosome. Male : XY ; Female : XX. Other 22 pairs of homologous chromosomes are called autosomes. The autosomes pairs are called homologous chromosomes. Human genome has above 3X10 9 bps in length. 97% of human genome is non-coding region called introns. 3% is responsible for controlling human genetic behaviour – exons.

4. Importance of genetics in medicine Almost 50% of first trimester abortion are due to chromosomal abnormalities. Congenital malformation : almost 2-3% of newborns. 2% infants are born with single gene disorder. More than half of childhood blindness, deafness and mental retardation are due to genetic disorder.

5. Salient features of double helix structures of DNA DNA : two polypeptide chains. Anti parallel polarity : 5’ to 3’ and 3’ to 5’. Backbone : sugar phosphate group and nitrogen base pair. Purine and Pyrimidine : A-T, G-C Coiled in a helical fashion. Out of 2 strands : one strand stores genetic information (template / anti-sense strand, 3` to 5’). It’s complementary strand : non-template / sense / coding, 5’ to 3’.

6. Structure of DNA Two polynucleotide strands. Two chains run in opposite directions. Sugar backbone. Base inside helix in complementary base pair. A = T ; C = G Strands bonded by weak hydrogen bonds.

7. Enzymes involved in DNA/RNA synthesis DNA/RNA polymerase : newly formed segment of DNA/RNA. Ligase : joining of segments. Endoneuclease : fragmentation.

8. Coiling of DNA Primary coiling of the duplex itself. Secondary coiling around histone beads. Tertiary coiling of nucleosome (chromatin fibres). Quarternary coiling forming loops.

9. Genome The total number of genes contained in one cell.

10. Genotype Complete genetic constitution of an individual.

11. Phenotype The physical appearance of an individual, that is, produced by the interaction of his genes and his environment.

12. Trait Variation of an expressed characteristics.

13. Karyotype Systemic arrangement of images of chromosomes into homologous pairs.

14. Allele Alternative forms of a gene that occupy a specific locus on a specific gene.

15. Homozygous Having two identical alleles for a given gene.

16. Heterozygous Having two different alleles for a given gene.

17. Carrier Heterozygous individual who does not display symptoms of a recessive genetic disorder but can transmit the disorder to his or her offspring.

18. Dominant Describes a trait that is expressed both in homozygous or heterozygous forms.

19. Recessive Describes a trait that is only expressed in homozygous form and is masked in heterozygous form.

20. X-linked Pattern of inheritance in which an allele is carried on the X chromosome of the 23 rd pair.

21. X-linked dominant Pattern of dominant inheritance that corresponds to a gene on the X chromosome of the 23 rd pair.

22. X-linked recessive Pattern of recessive inheritance that corresponds to a gene on the X chromosome of the 23 rd pair.

23. Transcription Copying of a DNA sequence into messenger RNA.

24. Translation Copying of messenger RNA into proteins.

25. Gene The unit of heredity that occupies a specific locus on the chromosome. It is a DNA sequence that directs synthesis of a specific polypeptide chain.

26. Types of genes Structural Regulatory Operator

27. Structural gene Directs synthesis of a specific polypeptide chain.

28. Regulatory gene Repressor substance secretion.

29. Operator gene Switches on structural gene.

30. Genetic code This is the language of messenger RNA based on the four bases of the m-RNA. “Words” are three RNA nucleotide sequences called codons.

31. Types of mutation Genetic mutation : results from changes in a single gene. Chromosomal mutation : involves changes in whole chromosome.

32. Gene mutation Change in the nucleotide sequence of DNA. Spontaneous mutation : molecular decay Induced mutation (mutagens) Chemicals Radiation Viral

33. Gene mutation occurs in 3 methods Inversion : base sequence reversed. GCC ( arg ) becomes CCG ( gly ). Substitution : replacement of one or more base pair with others. Transition : GTT to ATT (purine to purine) Transversion : ATT to CTT (purine to pyrimidine) Frame shift mutation : Insertion : addition of one or more nucleotide inside gene. Deletion : one or more nucleotide slipped out of gene.

34. Chromosomal mutation It occurs in 3 ways : Independent assortment of chromosome during meiosis : two pairs assort (2 2 = 4, 2 23 ) Random fertilisation Crossing over : during pachytene of meiosis between non-sister chromatids of homologous pair. Chromosomal mutation : in numbers Polyploidy Aneuploidy: Hyperploidy Hypoploidy

35. Intrachromosomal aberrations Deficiency : loss of terminal segment of chromosome due to single break. Deletion : loss of intercalary segment of chromosome due to double break in the chromosome followed by union of remaining part ( eg : ABCFGH / ABCDEFGH).

36. Interchromosomal aberrations Duplication : an extra chromosome segment attached to its normal homologous chromosome at its adjacent region of similar segment ( eg : ABCDCDEFGH / ABCDEFGH). Translocation : separation of chromosomal segment and its union to a non-homologous chromosome. Two types : Reciprocal : two non-homologous chromosome exchange segments between themselves to create new linkage (illegitimate crossing over). Robertsonian: one chromosome shows deletion or deficiency, while a non-homologous chromosome comes to have an additional segment (12, 13, 14, 21, 22 and Y).

37. Non-Mendelian inheritance Traits do not segregate in accordance with Mendel’s law. Mosaicism : 2 cell line present. Genetic imprinting Uniparental disomy Triplet repeat disorder : extra nuclear inheritance : Fragile X syndrome and Huntington’s disease. Genetic imprinting : between 2 copies of autosomal gene, one copy transcribed and other silenced. Methylation of DNA make one gene silent. 15q 11-12 gene (Prader Willi and Angelman syndrome). Microdeletion in paternal chromosome 15 leads to PW syndrome and maternal chromosome 15 leads to Angelman syndrome. UBE3A gene : responsible for AM syndrome. Microdeletion in maternal copy in brain resulting in insufficient protein production – leads to neurological manifestations. Uniparental disomy : inheriting both chromosome from mother – PW syndrome. Inheriting both chromosome from father – AM syndrome.

38. Mosaicism The presence of two or more types of cell population in the same individual. Post-zygotic mitotic non-disjunction will result in one trisomic and one monosomic daughter cell. The descendants of these cells will produce a mosaic.

39. Single gene mutation Deletion Duplication Inversion Insertion Translocation

40. Effects of gene mutation Neutral effect (silent mutation) Harmful effect Beneficial effect

41. DNA functions Replication : from one generation to another (inheritance). : from cell to cell (division). Metabolism : gene expression. : protein synthesis (transcription and translation).

42. Replication It is the process by which genetic information is transmitted to one cell to another. It starts by separation of the two strands. Each acts as a template for a new strand (semi – conservative). DNA polymerase proceeds in only one direction, along the helix (from 5’ to 3’ end of DNA), but starts in many sites.

43. Transcription The process whereby genetic information is transmitted from DNA to m-RNA by RNA polymerase into complementary single-stranded copies of RNA.

44. Codon A triplet of adjacent nucleotide in the m-RNA chain that codes for specific amino acid in the synthesis of a protein molecule.

45. Exons Nucleotide sequence that is found in a gene, code information for a protein synthesis and is transcribed to m-RNA.

46. Introns Segments of a gene situated in between exons that is removed before translation of m-RNA and does not function in coding for protein synthesis.

47. Promoter The place where RNA polymerase first attaches. They vary from gene to another but contains sequences in common of seven base sequences of all T’s and A’s.

48. Terminator Certain sequences that terminates gene transcription (downstream beyond the 5’end).

49. Sense strand The strand that is actually transcribed is the one with specific promoter sequence. The other non-sense strand is not used in transcription.

50. Processing Capping : addition of a chemical cap to 5’ end (attach RNA to ribosome). Splicing : excision of introns. Polytail : insertion of poly-A tail (AAAA) to the 3’ end (aids in transporting RNA molecule to cytoplasm).

51. Translation It is the process by which m-RNA is translated into protein synthesis. It involves t-RNA and r-RNA.

52. t-RNA One end binds to a particular amino acid (all are 20), the other ends has a 3 base sequence, the anti-codon, that pairs with a particular codon on m-RNA.

53. r-RNA Structural elements of ribosome. They are the benches on which protein synthesis occurs.

54. Pattern of inheritance Autosomal recessive Autosomal dominant X-linked recessive X-linked dominant

55. Autosomal recessive Heterozygotes are carriers with normal phenotype. Most affected children have 2 normal parents (Aa X Aa). 2 affected parents will always produce affected children ( aa X aa ). Both males and females affected equally. Pedigree shows male and female carriers.

56. Autosomal recessive

57. Autosomal dominant Heterozygotes are affected. Affected children usually have affected parents. 2 affected parents can produce an unaffected child (Aa X Aa). 2 unaffected parents will not produce affected child. Both males and females affected equally. Male to male transmission that eliminate X-linked inheritance. Pedigree shows no carrier.

58. Autosomal dominant

59. X-linked recessive More males than females are affected. Female carrier have 25% chance of having an affected son, 25% carrier daughter and 50% normal. Affected males have carrier daughter and unaffected son. Male to male transmission excludes X-linked but is seen in autosomal dominant and Y-linked inheritance.

60. X-linked recessive

61. X-linked dominant If father affected and mother unaffected, there is probabilities of 100% affected daughter and all sons are normal. Unaffected father and affected mother will produce 50% affected son and 50% affected daughters. No carriers.

62. X-linked dominant

63. Y-linked inheritance Very rare. Only male to male transmission. Most of the cases are infertile (sex determination site is involved). Leri- weil dyschondrosteosis. Pseudo autosomal sites are affected.

64. Numeric abnormalities of chromosomes Normal chromosomal number is 46. it is called euploid. Polyploidy : more than 2 sets of homologous chromosomal. Chromosomal number is 3n or 4n. Generally results in spontaneous abortion. Aneuploidy : any chromosomal number that is not an exact multiple of haploid number., that is 47 or 45.

65. Aneuploidy Most common cause is non-disjunction of either a pair of homologous chromosomes during meiosis I or failure of sister chromatids to separate during meiosis II. The result gamete will have either one less chromosome or one extra chromosome. Fertilisation of such gamete will result in zygote being either trisomic or monosomic. Monosomy in autosomes is incompatible with life. Trisomy of certain autosomes and monosomy of sex chromosomes is compatible with life.

66. Human genome project (HGP) Started in 1 st October 1990, under the department of Energy and National Institute of Health in USA. Later, UK, France, Japan, Germany, China and few other countries joined. The project was completed in 2003. This was a 13 year project.

67. HGP is a mega project Huge cost (9 billion USD). Very large number of bases pairs to be identified and sequenced (3X10^9). Storage of data generated which requires 3,300 books, each with 1000 pages, each having 1000 typed letters. High speed computational device for storage, retrieval and analysis of data made it easier.

68. Goals of HGP Identification of 25,000 genes in human DNA. The number of gene is still not known precisely. To determine the sequence of 3 billion chemical base pairs. Most of the human genome does not encode protein. Less than 5% being transcribed and translated. To store the information in database. To improve tools for data analysis. To resolve any ethical, legal and social issues (ELSI) that may arise from the project. Sequencing of model organism.

69. Salient features of human genome Human genome contains 3 billion nucleotide base. The average gene consists of 3,000 bases, but their size varies greatly. The size of genome or number of gene connected with the complexity of thee body organism. The functions of 50% of discovered gene is unknown. By differential splicing, a gene may code for two or more proteins. Chromosome number 1 has maximum number of gene (almost 3,000) while Y chromosome has minimum number of genes (231). 99.9% of nucleotide bases are exactly similar in all human beings. Part of DNA which contains repeated sequence is called satellite DNA. Single nucleotide polymorphism (SNPS) have been identified in humans, their number maybe more than 10 million. This will be helpful in finding out the chromosomal location with disease associated sequence and tracing human history.