Spectrum biology

TATA Box

The TATA box of the DNA is the site of assembly of a preinitiation

complex that contains the general transcription factors and the polymerase.

The TATA box within eukaryotic promotors include

(a)Nucleotide sequence of the region just upstream from the site where

transcription is initiated in three different eukaryotic genes. The TATA box is

indicated by the grey shading.

(b)Three-dimensional structure of the TATA Binding Protein (TBP) from the

flowering plant Arabidopsis thaliana as it sits astride the DNA.

(c)Binding of TBP is accompanied by a dramatic distortion in the conformation of

the DNA helix, which unwinds over an eight base pair stretch.

Genetic Code is not always Universal: Do you

know?

In few organisms, some of the codons code uniquely different from rest of the

organisms (universal). Some of them are tabulated below.

Codons*

AGA

UGA AUA AGG CUN CGG

Normal code assignment Stop lle Arg Leu Arg

Vertebrates Trp Met Stop + +

Yeasts (Saccharomyces

cerevisiae)

Trp Met + Thr +

Filamentous fungi Trp + + + +

Higher plants + + + + Trp

Chlamydomonas reinhardtii? + + +?

*N indicates any nucleotide; + codon has the same meaning as in the normal

code?, codon not observed in this mitochondrial genome.

Hu man Ge nome Pro ject (HGP)

HGP is a comprehensive international research effort dedicated to
map the entire genome by determining the sequence of nucleotides
in the DNA of each of the 22 +X and Y-chromosomes and to study
the functions of human genes. It has also been called International
Human Genome Sequencing Consortium.

Goals of HGP

(i)To determine the sequence of the 3 billion base pairs present in

the human genome.

(ii)To identify and determine the functions of approximately all

20000-25000 genes in human DNA.

(iii)To improve tools for data analysis.

(iv)To transfer the technologies to other sectors, such

as industries.

(v)To address the ethical, legal and social

implications of the results obtained from the

project.

(vi)To identify thousands of genetic markers and map

them in the genome.

Strategy and Methodology

Methodology of HGP includes following three stages:

1.Mapping The physical and genetic maps of

human genome are prepared by using molecular

markers, microsatellites, simple sequence repeats

or sequence tagged sites and PCR amplification of

particular microsatellites.

2.Sequencing The method employed for

sequencing the entire genomic DNA of human

have two basic approach.

3.Generation of physical and genetic maps:

The genetic and physical maps of genome were

generated by using information on:

(i)Polymorphism of restriction endonuclease

recognition sites.

(ii)Some repetitive DNA sequences called

microsatellites.

The HGP Strategy

Clones isolated from a genomic library were ordered

into a detailed physical map, then individual clones

were sequenced by shotgun sequencing protocols.

The strategy used by the commercial sequencing effort

eliminated the step of creating the physical map and

sequenced the entire genome by shotgun cloning.

It clearly showed that only 1.1%-1.4% part of human

genome code for proteins.

Sequence annotation

It involves determining the

complete genome sequence

including all the coding and

non-coding sequences and

then assigning functions to

different regions in the sequence.

Sequencing

Expressed sequence

tagging method

This method involves

identifying all those genes

that are expressed as RNA.

This is represented as ESTs

Chicken

ovalbumin

Rabbit

β-globin

Mouse

β-globin

major

GAGGCTATATATTCCCCAGGGCTCAGCCAGTGTCTGT CAA

TTGGGCATAAAAGGCAGAGCAGGGCAGTGCTGCTA CACTA

GAGCATATAAGGTGAGGTAGGATCAGTTGCTCCTC CATTTA

(a)

(b)

(c)

NH 2

COOH

TBP

DNA

TATAAATA

DNA is digested into fragments; fragments

inserted into Bacterial Artificial Chromosomes (BAC)

Fragments are identified and mapped.

BAC to be sequenced is fragmented;

fragments sequenced at random.

Sequence overlaps reveal final sequence.

Final sequence

Genomic DNA