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INTRODUCTION
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It involves the fusion of somatic cells growing in vitro.
First demonstrated with mouse cells by G.Barski
(1960).
Somatic cell hybrids can be produced between two cell
of same species or two cells from different species. For
ex. Mouse-human hybrids.
Used for mapping human gene.
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Human – mouse hybrids are usually made using
established mouse cell culture lines and human
fibrocytes or leukocytes.
 Important features of human-mouse hybrids:1. Mouse-human chromosomes are easily
distinguished.
2.Both sets of chromosomes of human and mouse
are expressed in hybrid cells.
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involves fusion of somatic cells
growing in vitro.
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Genes are shown to be located
on specific chromosomes by
correlating the phenotypes ex.
Enzyme activities controlled by
these genes with presence of
individual human chromosomes
in hybrid cell clones. For
example, human cell make
enzyme thymidine kinase (TK+)
were hybridized with mouse
cell make enzyme hypoxanthineguanine
phosphoribosyltransferase
(HPRT).
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Frequency of spontaneous fusion
is very low - increased by
addition of ultraviolet-inactivated
sendai virus or chemical
polyethylene glycol- agent
stimulate cell fusion by
increasing cell contact and
altering cell membrane.
Cell fusion produces binucleate
hybrid cells or heterokaryons.
Cell fusion is followed by
nuclear fusion to produce
uninucleate hybrid cell or
synkaryons
hybrid cell clones will grow on
HAT medium.
Each of 24 different
chromosomes can identified
using G-banding.
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BIOCHEMICAL BASIS OF SELECTION PROCEDEURE
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Involves two important pathways for biosynthesis
of nucleotides require for DNA synthesis i.e. de
novo pathway and salvage pathway.
de novo pathway : major pathway by which
nucleotides are synthesized from simple sugars and
amino acids or this pathway is blocked by
antimetabolite aminopterin.
If de novo pathway is blocked, then second
pathway called salvage pathway comes.
Salvage pathway: HGPRT catalyzes the conversion of
hypoxanthine to guanine and thymidine kinase catalyzes the
phosphorlyation of thymidine.
Enzyme TK+ and HGPRT+ are necessary for salvage
pathway.
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Recombination fraction is measure of distance between the two loci.
oTwo genes that undergo independent assortment have recombination
frequency of 50 percent and are located on non homologous
chromosome or far apart on same chromosome – unlinked.
oGenes with recombination frequencies less than 50 percent are on
same chromosome –linked.
o
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TO CALCULATE RECOMBINANT FREQUENCY:
The Recombination frequency for interval from a
gene to centromere is one-half the frequency of asci
that exhibit second-division segregation patterns for
the alleles of the gene.
For example: