Gene editing methods allow for targeted modification of DNA in living organisms. The document outlines the basic steps of gene editing which include using an engineered nuclease to induce double-stranded breaks in target DNA, followed by repair of the breaks. It discusses three main gene editing technologies - ZFN, TALEN, and CRISPR/Cas9 - and explains how CRISPR/Cas9 has been applied successfully to modify crops. The document concludes that CRISPR-based gene editing has become an efficient and flexible tool for plant research and crop improvement.

1. Triticeae Research Institute, Sichuan Agricultural University
四川农业大学小麦研究所
Course: Advanced Plant Molecular Biology
Presentation Topic: Gene Editing
Supervisor: Prof. Dr. Qi Pengfei
Name: Muhammad Sajid Sharif

2. Steps in Gene Editing
CONTENTS
01
02
03
05
Introduction
Background of Gene Editing
CRISPR/Cas9 and crops
04 Types of G.E
06 Conclusion

3. 1 Introduction

4. Gene Editing
Introduction
01
Gene-editing methods have arisen as an efficient tool
for rapid analysis of gene function (Kumar et al,
Plant biotechnology Journal, 2018.)
Gene Editing is a type of engineering in which
DNA is inserted, deleted ,modified or replaced in
living organism including the crop plant.

5. 2 Background of Gene
Editing

6. Production of plant with improved traits
desireable. Traditionally plant breeding method
or labor and time consuming . Gene Editing
allows targating and modifying specific DNA
sequences.
Background of Gene Editing
02

7. 3 Steps of Gene Editing

8. Steps of Gene Editing
03
• First exogenous engeried nucleaous
containing recogination module and
nucleause recognize the target DNA.
• Then engeried nucleus bind to target
DNA and induces double Stroke
breaks (DSBs).
• The Homology - directed Repair (HDR)
pathway then repair the (DSBs).
• HDR result in repair of (DSBs).

10. 4 Types of G. E.

11. Types of Gene Editing
04
There are three types of Gene Editing:
1.Zinc fingr nuclease (ZFN)
2.Transcription activator like effecetor
nuclease (TALEN)
3.Clustered Regularly interspaced short
palindromic repeat (CRISPR)

12. ZFNs and TALENs consist of
sequences specific DNA binding
module and a Fokl nuclease domain .
The Fokl nuclease domain requires
damerization to become an active
nuclease .

13. 5 CRISPR/Cas9 and Crops

14. CRISPR/Cas9 and Crops
05

15. CRISPR/Cas9 and Crops
05
The CRISPR system emerged as gnome editing tool in 2012.
Components of CRISPR
There are two majors components :
I. Cas9 Protein
II. Guide RNA (gRNA)
Cas9 Protein is a RNA depenedent DNA endo nuclease
that form a complex with (gRNA)

16. Gene Editing Using CRISPR/ Cas9 in Plants
05
The CRISPR system has been successfully applied in various plant species.
These include not only model plants such as arabidopsis,but also crops
such as rice, tobacco, sorghum,wheat,maze,soyabean,tomato,apple and
banana.
The purpose of this application includes the enhancement of abiotic and
biotic stress resistence, engineering of metabolic pathways and increase
green yeild.
The introduced mutation are inherited by next genration of plant indicating
that plant gneome editing can be used for plant research and production
of usefull plants.

17. WhatCRISPR and How it works?
CRISPR molecule is made of short palindromic DNA
sequeces that are repaeted along the molecules and a
regularly between the spaces forgiven DNA sequnce
from the orgsnism that have previously attacked the
bactaria. The CRISPR molecule also includes CRISPR
associated genes. These encodes protiens that unwind
DNA , and cut DNA .

18. CRISPR Allows Reseacher to Perform the Following
CRISPR allow researchers to
perform the following
1.Gene Knock-out
2.DNA- Free Gene Editing
3.Gene Insertions
4.Transient Gene Silencing

19. Gene Knock -Out
05
Gene silencing using CRISPR starts with the use of a
single guide RNA to target genes and initiate a double
standered break using Cas9 endo nuclease .
DNA-free Gene Editing :
CRISPR can be used for DNA-free gene editing without the
use of DNA vectors, requiring only RNA or protien
componentes.A DNA-free gene editing system can be good
choice to avoide the possibillty of undesireable genetic
alterations due to the plasmid DNA integrating at the cut site
or random vectoe integrations.

20. Gene insertions
The CRISPR induced double stand break can also be used to create gene
“knocks -ins” by exploitaing the cells homology directed repair. The precise
insertion of a donner templete can alter the coding region of a gene.
Ttransient Ggene Silencing:
By modifying the Cas9 protien so it cannot cut DNA
transient gene silencing can also be done .The modyfied
cas9 oled by a guide RNA targets the permoter region of
genes and reduce transcriptional activity and gene
expression.

23.  With the emergenc of new CRISPR based targeted gene
editing has become increasely efficient an flexible in
the plant cell
 In addition to targeted gene mutagensis other types of
gentic modifications have become achievable in
multiple plant species
Conclusion Remarks :

24. THANKS for your Pationace