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Bc 100 Dna Recombination

DNA recombination is the process of breaking and rejoining parental DNA molecules to create new genetic combinations, playing crucial roles in gene diversity, DNA repair, and the production of transgenic organisms. There are several types of recombination, including homologous, nonhomologous, site-specific, and replicative recombination, each serving different biological functions. Meiotic recombination enhances genetic variation in offspring, essential for sexual reproduction, and involves complex mechanisms such as the Holliday model and site-specific recombination.

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Mohammed Sibghatullah Haidar
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20 views30 pages

Bc 100 Dna Recombination

DNA recombination is the process of breaking and rejoining parental DNA molecules to create new genetic combinations, playing crucial roles in gene diversity, DNA repair, and the production of transgenic organisms. There are several types of recombination, including homologous, nonhomologous, site-specific, and replicative recombination, each serving different biological functions. Meiotic recombination enhances genetic variation in offspring, essential for sexual reproduction, and involves complex mechanisms such as the Holliday model and site-specific recombination.

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Mohammed Sibghatullah Haidar
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DNA RECOMBINATION

DR F DIDA. MD, PhD


Definition of recombination
• Breaking and rejoining of two parental DNA
molecules to produce new DNA molecules
Biological Roles for Recombination

1. Generating new gene/allele combinations


(crossing over during meiosis)
2. Generating new genes (e.g., Immuno-
globulin rearrangement)
3. Integration of a specific DNA element
4. DNA repair
Practical Uses of Recombination
1. Used to map genes on chromosomes

2. Making transgenic cells and organisms


Types of Recombination

• At least four types of naturally occurring


recombination have been identified in
living
organisms
• General or homologous recombination
occurs between DNA molecules
of very similar sequence, such as homologous
chromosomes in diploid organisms. General
recombination can occur throughout the genome of
diploid organisms, using one or a small number
of common enzymatic pathways.
• Illegitimate or nonhomologous
recombination
occurs in regions where no large scale
sequence similarity is apparent, e.g.
translocations between different
chromosomes
• Site-specific recombination occurs
between particular short sequences (about 12 to 24
bp) present on otherwise dissimilar parental
molecules. Site-specific recombination requires a
special enzymatic machinery, basically one
enzyme or enzyme system for each particular site.
Good examples are the systems for integration
of some bacteriophage, such as λ, into a bacterial
chromosome(DNA recombinat technique) and the
rearrangement of
immunoglobulin genes in vertebrate animals.
• replicative recombination, which
• generates a new copy of a segment of DNA. Many
transposable elements (Transposable elements
(TEs), also known as "jumping genes" or
transposons, are sequences of DNA that move
(or jump) from one location in the genome to
another. use a process of
• replicative recombination to generate a new copy
of the transposable element at a new location.
Types of recombination
A+ B+ C+ A+ B- C-
Homologous
or general
A- B- C- A- B+ C+

Nonhomologous A B C A B F
or illigitimate
D E F D E C

 att  integrase

Site-specific
att att
att E. coli

Replicative transposase
recombination, A B C A B C
transposition
Transposable element
Recombination
• Breaking and rejoining of two parental DNA
molecules to produce new DNA molecules
• Reciprocal recombination: new DNA
molecules carry genetic information from
both parental molecules.
• Gene conversion: one way transfer of
information, resulting in an allele on one
parental chromosome being changed to the
allele from the other homologous
chromosome
Gene Conversion

A+ B+ C+ A+ B+ C+

A- B- C- A- B+ C-
• Homologous or general recombination:
– Bacterium with two viruses (transformation:
refers to the introduction of foreign DNA into
bacterial)
– Bacterium after conjugal transfer of part of a
chromosome (transfection: refers to the
introduction of foreign DNA into mammalian
cells)
– At chiasmata during meiosis of eukaryotic cells
– Post-replication repair via retrieval system
B. Meiotic recombination
• Recombination appears to be needed to keep maternal
and paternal homologs of chromosomes together prior to
anaphase of meiosis I
– Zygotene: Pairing of maternal and paternal chromosomes (each has 2
sister chromatids)
– Pachytene: Crossing over between maternal and paternal chromosomes
– Diplotene: Centromeres of maternal and paternal chromosomes separate,
but chromosomes are held together at chiasmata (cross-overs)
– Anaphase I: Homologous chromosomes separate and move to 2 daughter
cells.
• Results in >1 exchange between pairs of homologous
chromosomes in each meiosis.
• Failure to keep homologous chromosomes together prior to
anaphase I can lead to aberrant numbers of chromosomes,
e.g. trisomy for chromosomes 15, 18, 21
Cross-overs during meiosis I
Zygotene: Homologous
chromosomes,
Maternal
each with 2 sister chromatids, pair Paternal
to form bivalents (line=duplex DNA)

Pachytene: Cross-overs between


homologous chromosomes

Diplotene: homologous
chromosomes separate partially but
are held together at cross-overs
Metaphase I
Anaphase I
Anaphase I: Cross-overs resolve
to allow homologous chromosomes
to separate into separate cells

Meiosis II
Benefits of recombination
• Greater variety in offspring: Generates new
combinations of alleles
• Negative selection can remove deleterious
alleles from a population without removing
the entire chromosome carrying that allele
• Essential to the physical process of meiosis,
and hence sexual reproduction
Meiotic recombination generates new
combinations of alleles in offspring
Each line is duplex DNA, starting at pachytene of meiosis I

A1 B2 C2 A1 B3 C1
Dad Mom
A2 B1 C4 A3 B1 C3
Finish Meiosis I
Meiosis II
A1 B2 C2 A1 B3 C1
A1 B2 C4 A3 B3 C1
A2 B1 C2 A1 B1 C3
A2 B1 C4 A3 B1 C3
Fertilization

A1 B2 C4
A3 B3 C1 Child
Holliday model for recombination
• Pairing: align homologous duplexes
• Single strand invasion:
– Endonuclease nicks at corresponding regions of
the same strands of homologous chromosomes
– Ends generated by the nicks invade the other,
homologous duplex
– Ligase seals nicks to form a joint molecule.
– (“Holliday intermediate” or “Chi structure”)
• Branch migration expands heteroduplex
region.
Holliday Model
R. Holliday (1964)

- Holliday Junctions
form during
recombination

- HJs can be resolved


2 ways

patch
EM of a Holliday Junction w/a few melted
base pairs around junction

Fig. 22.3
Resolution of joint molecules
• Can occur in one of two ways
• The Holliday junction can be nicked in the
same strands that were initially nicked =
“horizontal resolution.” This results in NO
recombination of flanking markers.
• The Holliday junction can be nicked in the
strands that were not initially nicked =
“vertical resolution.” This results in
RECOMBINATION of flanking markers.
Vertical & horizontal resolution
A+

B+
V

B-
A-

or 1. Horizontally H
2. Vertically V
A+ B+
A+ B-

A- B-
A- B+

This leaves a region of heteroduplex, and A region of heteroduplex is left, but the
the flanking markers have recombined. flanking markers are not recombined.
Common steps in models

• Generate a single-stranded end


• Search for homology
• Strand invasion to form a joint molecule
• Branch migration
• Resolution
Site-specific recombination
• Moves specialized nucleotide sequence (mobile
genetic elements) between non-homologous sites
within a genome.
• Transpositional site-specific recombination
• Conservative site-specific recombinatinon
Transpositional site-specific
recombination
• Modest target site selectivity and insert mobile
genetic elements into many sites
• Transposase enzyme cuts out mobile genetic
elements and insert them into specific sites.
Cut and Paste Transposition
DNA-only
Replicative Transposition
Retrovirus-based Transposition
Retroviral-like retrotransposition
Reverse Transcriptase

RNA

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