rnai technology

Presented By:- Dongare P.B.7th sem

BED11ABT03B

Guided By:-Prof. Pawar S.S.

RNA interfernce (RNAi) is an evolutionally highly

conserved process of post-transciptional gene

silencing (PTGS) by which double stranded RNA

(dsRNA) cause sequence specific degradation of

mRNA sequence.

It was first discovered in 1998 by Andrew Fire and

Craig Mello in the nematode worm and later found in

a wide variety of organisms, including mammals.

PTGS (Post-transcriptional Gene Silencing)

Cosuppression

Quelling

Virus-induced gene silencing

The discovery of RNAi was first observed by Antisense RNA expressed in transgenic plant by plant scientist in United state and Netherland in the early 1990.

RNAi first observed in petunia plant flower.

Petunia plant in which genes for

pigmentation are silenced by RNAi

Craig Mello, born in 1960, is

a US citizen and professor of

molecular medicine.

Andrew Fire, born in 1959, is

a US citizen. Since 2003 he

has been professor of

Pathology and Genetics at

Stanford University.

Fire and Mello made their key

discoveries about RNA

Interferance.

Craig Mello

Andrew Fire

Gene silencing

mediated by double-

stranded RNA.

RNA-dependant gene

silencing process that

is controlled by the

RNA-induced silencing

complex(RISC). Mechanism of RNA

interfance in Mammalian

cells.

1. Endogenous dsRNa initiates RNAi by activating the

ribonuclease protein Dicer.

2. It is bind and cleave dsRNA to produce double

stranded fragment of 20-25 bp.

3. Double stranded RNA triggers peocessed in siRNA

by enzyme RNAseIII family, specifically the Dicer

family.

4. Dicer family proteins are ATP-dependant

nucleases.

These protein contain an Amino-terminal helicasedomain.

Dicer homologous exist in many organism including c.elegance, Drosophilla, yeast and humans.

Developmental consequence in Drosophilla and c.elegance.

The dicer protein

which catalyzes the

cleavage of dsRNA

to siRNAs

> RISC is large (500-kDa) RNA-multiproteincomplex, which triggers mRNA degradation in Response to siRNA.

> Some componants have been defined by Genetics, but function is unknown.

> Cleaved mRNA is degraded by cellular exonucleases.

The active components of an RNA-induced silencing complex (RISC) and endonucleasescalled argonaute proteins, which cleave the target mRNA strand complementary to their bound siRNA.

A Full-length argonaute protein from the

archaea species.

1. RNA-induced transcriptional

silencing (RITS), and is

carried out by a complex of

protein called the RITS

complex.

2.The mechanism by which the

RITS complex induced

heterochromatin formation

and orgnization is not well

understood.

3. In maintenance of existing heterochromatin

regions, RITS forms a complex with siRNA

complementary to the local genes.

4. The formation of such a heterochromatin region

is dicer-dependant because dicer is required to

generate the initial complement of siRNA.

1. Alteration of plant aechitecture

2. Abiotic and Biotic stress tolarance.

3. Nutritional Improvement.

4. Deletion of Allergenes.

5. Removal of toxic compounds.

6. Prolongation of shelf life.

7. Modulation of Flower colour and fragrance.

8. Engineering of secondary Metabolites.

9. Seedless fruit development.

10. Development of Male sterile plants.