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OR OR What is lineage tracing Schematic of a late embryo/adult Schematic of an early embryo Schematic of an early embryo Schematic of an early embryo Schematic of a late embryo/adult
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Lineage tracing and fate mapping 1
BSE652 Lineage tracingandfate mapping 1 Lecture 2 7th Jan 2016 OR
OR What is lineage tracing Schematic of a late embryo/adult
Schematic of an early embryo Schematic of an early embryo Schematic
of an early embryo Schematic of a late embryo/adult What is a Fate
map? This chart or graphical representation detailing the fate of
each part of an early embryo in the adult animal is referred to as
a fate map At a single cell level fate mapping is known as lineage
tracing
Why is it important to learn the lineage of a cell or fate map of
an early embryo? To know the ground truth To understand the
journey, change in molecular signature and microenvironment, of a
differentiated cell To affect specific molecular manipulations in
specific cell populations Methods of lineage tracing
Direct observation Methods of lineage tracing
Conklin (1905) - Tunicates Methods of lineage tracing
Direct observation Conklin (1905) - Tunicates Direct observation,
time lapse video microscopy followed by cell ablation Limitations:
This method currently gives the best resolution in space and time,
but has the disadvantage that the number of cells which can be
followed in a single individual is limited by the short-term memory
of the observer. Cell number and opaque embryo DYE LABELING
Schematic showing agar chips with vital dyes applied onto the
surface of an early stage amphibian embryo (top). These dyes label
regions within later stage embryos (bottom) (based on Vogt, 1929;
adapted from Gilbert, 2000). Use of lipophilic dye conjugated with
large molecules (dextran) Taking advantage of cell proliferation
properties EdU First observation made in chicken, Followed up in
mice EdU , Day10 EdU pulse chase to lineage trace proliferative
cells
EdU injected intraperitoneally with 100 mg/kg body weight into
13.5/15.5 dpc pregnant females EdU 2 Hour So, we wanted to lineage
trace the proliferative population of cells. For this purpose we
injected EdU in pregnant females with to label proliferating cells
in the skeletal tissues. In the first set, In this experiment, two
sets of mice were taken; in the first set around EdU was injected
and chased for 2 hours. In the second set EdU injected was chased
for 24 hours. The data are as follows. EdU pulse chase to lineage
trace proliferative cells
So, we wanted to lineage trace the proliferative population of
cells. For this purpose we injected EdU in pregnant females with to
label proliferating cells in the skeletal tissues. In the first
set, In this experiment, two sets of mice were taken; in the first
set around EdU was injected and chased for 2 hours. In the second
set EdU injected was chased for 24 hours. The data are as follows.
EdU 2 hour pulse EdU pulse chase to lineage trace proliferative
cells
EdU injected intraperitoneally with 100 mg/kg body weight into
13.5/15.5 dpc pregnant females EdU 2 Hour So, we wanted to lineage
trace the proliferative population of cells. For this purpose we
injected EdU in pregnant females with to label proliferating cells
in the skeletal tissues. In the first set, In this experiment, two
sets of mice were taken; in the first set around EdU was injected
and chased for 2 hours. In the second set EdU injected was chased
for 24 hours. The data are as follows. EdU 24 Hour Distal
Proliferative Zone (DPZ) cells indeed contribute to the growth of
interzone
EdU 24 hour pulse chase EdU 2 hour pulse Slowly proliferating
interzone cells do not account for the EdU incorporation after 24
hours DPZ cells contribute to articular cartilage Retinal
development and differentiation Retroviral infection The neural
retina has seven different cell types: six neuronal and one
glial
These cells are organized into three nuclear layers: GCL (Ganglion
cell layer), containing the cell bodies of the ganglion cells. INL
(Inner nuclear layer) containing the cell bodies of the bipolar
cells, horizontal cells and amacrine cells. ONL (outer nuclear
layer), containing the nuclei of the photoreceptors e.g. Rods and
cones. In addition to these nuclear layers there are two plexiform
layers: IPL (inner plexiform layer), containing the synapses and
processes of the bipolar, amacrine and ganglion cells. OPL (inner
plexiform layer), containing the synapses and processes of the
photoreceptor, bipolar and horizontalcells. The cell types are
produced in an orderly manner which is generally conserved among
vertebrates.
Ganglion cells are produced first. Followed closely by cone,
horizontal and amacrine cells Then come rod and bipolar cells The
cell type to be born are the Muller glia cells. During
retinogenesis these seven cell types derive from a common
population of retinal progenitor. Lineage analyses has revealed
that retinal progenitors are multipotent and retain their ability
to generate different cell types up to the final cell division. If
an individual retinal progenitor is injected with a genetic marker,
that marker will be seen in a strip that includes all different
cell types in the retina. Determination of the lineage of a
neuroblast in the rat retina. (A) Technique whereby a virus
containing a functional -galactosidase gene is injected into the
back of the eye of a newborn rat to infect some of the retinal
precursor cells. After a month to 6 weeks, the eye is removed and
the retina is stained for the presence of -galactosidase. (B)
Stained cells forming a strip across the neural retina, including
five rods (r), a bipolar neuron (bp), a rod terminal (t), and a
Mller glial cell (mg). To distinguish between the possibilities of
(1) the same progenitor cell gives rise to two different daughters
Vs (2) two closely spaced DISTINCT cells give rise to distinct
daughters This study was made even more comprehensive by
co-relating with cell division Another method of permanent
labelling of cells: Quail-Chick Chimeras
Methods in Molecular Biology Volume 97, 1999, pp Quail-Chick
Chimeras Marie-Aime Teillet, Catherine Ziller, Nicole M. Le
Douarin