What is gene targeting in plants?

More specifically, gene targeting is the alteration of a specific DNA sequence in an endogenous gene at its original locus in the genome, and often refers to the conversion of the endogenous gene into a designed sequence [2]. Double-stranded breaks in plant genomic DNA are repaired either via HR or NHEJ [7].

What are the different methods of gene transfer in plants?

Method # I. Vector-Mediated Gene Transfer:

There are mainly two species of Agrobacterium: i.
Crown Gall Disease and Ti Plasmid:
Organization of Ti plasmid:
T-DNA region:
Virulence region:
Opine catabolism region:
T-DNA transfer and integration:
Signal induction to Agrobacterium:

What is targeted gene transfer?

Gene targeting (also, replacement strategy based on homologous recombination) is a genetic technique that uses homologous recombination to modify an endogenous gene. The method can be used to delete a gene, remove exons, add a gene and modify individual base pairs (introduce point mutations).

What is gene transfer in plants Slideshare?

3. INTRODUCTION • Gene transfer or uptake of DNA refers to the process that moves a specific piece of DNA into cell. • The directed desirable gene transfer from one organism to another and the subsequent stable integration & expression of foreign gene into the genome is referred as genetic transformation.

What is a targeting construct?

The targeting construct is usually a plasmid that contains two long stretches of genomic DNA, called homology arms, which are designed to match as closely as possible the genomic DNA of the ES cell line being targeted.

What is the role of the target gene?

“Target Gene” often just means “Gene Of Interest”, or the particular gene being studied or manipulated in an experiment. In the context of a “gene knockout”, a “target gene” may be the gene that a “targeting vector” is designed to knock out (make non-functional, non-stable, or non-expressable).

What are the two methods of gene transfer?

The methods used for gene transfer fall into two main categories; natural and artificial transformation.

Which is the direct method of gene transfer?

Complete answer: 1. Microinjection: In this method of direct gene transfer, the desired DNA segment is injected directly into plant protoplasts and cells (the nucleus or cytoplasm), this is done using fine-tipped glass needles. This glass needle is also referred to as a micropipette.

What is gene transfer method?

The artificial gene transformation can also be conducted through chemical methods which include calcium phosphate-mediated, polyethylene glycol-mediated, DEAE-Dextran, and liposome-mediated transfers. Electrical methods are also artificial ways to transfer genes that can be done by electroporation and electrofusion.

What is direct gene transfer?

The term direct transfer of gene is used when the foreign DNA is directly introduced into the plant genome. Direct DNA transfer methods rely on the delivery of naked DNA into the plant cells.

What is a gene targeting vector?

Targeting vectors are designed with the selection marker placed in an intron flanked by loxP or FRT recognition sites. The endogenous exon is then replaced with the mutated exon/intronic marker and the selection marker is subsequently deleted from the genome via site-specific DNA recombination.

What is the structure and function of genes?

Gene structure is the organisation of specialised sequence elements within a gene. Genes contain the information necessary for living cells to survive and reproduce. In most organisms, genes are made of DNA, where the particular DNA sequence determines the function of the gene.

Which is the target line for PLANTA gene targeting?

The target/donor combination line T-13/D-28, in which both constructs are located on chromosome I, was selected, and its F2′ generation was screened for completely stained plants ( Fig. 2 B ).

How are site specific endonucleases used in plants?

The development of designed site-specific endonucleases boosted the establishment of gene targeting (GT) techniques in a row of different species. However, the methods described in plants require a highly efficient transformation and regeneration procedure and, therefore, can be applied to very few species.

How is the progeny of a targeted allele obtained?

Progeny clonal for the targeted allele could be obtained directly by harvesting seeds. Targeted events could be identified up to approximately once per 100 seeds depending on the target donor combination. Molecular analysis demonstrated that, in almost all events, homologous recombination occurred at both ends of the break.

How big is a fragment of Planta gene?

Whereas a 6.1-kb fragment is indicative of the native target locus (T-13) and a 3.6-kb fragment of the donor construct (D-28), integration of the GT vector via HR would result in a 5.6-kb fragment. In all 20 tested lines, such a 5.6-kb fragment could be detected (see GT-1 to GT-6).

What is gene targeting in plants? More specifically, gene targeting is the alteration of a specific DNA sequence in an endogenous gene at its original locus in the genome, and often refers to the conversion of the endogenous gene into a designed sequence [2]. Double-stranded breaks in plant genomic DNA are repaired either via HR…