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by Miguel
A. Germain
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The processes by which an
organism develop from a single cell seems miraculous,
and there is no more exciting
event than that of birth. The adaptation of the
the newborn to it new life
is also exhilarating to witness.
GERMANIUM
The purpose of this activity is to encourage biology ,
marine biology , anatomy and physiology students to investigate the effects
of mutagen and teratogen in the development of zebrafish .
As natural mutation is a biological affect of evolution, any substance
that increases the rate of gene mutation may also act as a carcinogen.
This activity will further the students understanding of the mutation process
that occur with the DNA strand ,as well as the process by which a
mutation is translated from DNA into the protein that makes up the organism's
structure. One of the most important skills that students should develop
during any laboratory science course is the ability to use the scientific
method of experimentation.
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This activity is not designed to duplicate
scientific experiment conducted by researchers but rather to complement
and enhance students learning. The purpose of this activity is to
give students the ability to design other laboratory experiments.
There are numerous variable that the students and teachers can undertake.
In this investigation students will view morphological
changes induce by chemicals into developing zebrafish eggs.
Congenital malformations are
anatomical abnormalities are present at birth. They may be macroscopic
on the surface of the body or microscopic within the body.
About 20 per cent of deaths in the perinatal period are attributed
to congenital malformation (Mac Vicar,1976). Malformation
are observed in about 2.7 per cent
of newborn infants. Congenital abnormalities are detected
in an additional 3 per cent (Mc Keown, 1976).
Genetic factors initiate mechanism of malformation by
biochemical or other means at the subcellular, cellular, or tissue level.
The mechanism initiated by the genetic factor may be identical with or
similar to the causal mechanism initiated by teratogen or mutagen.
The simplicity of these activities
can be performed by students in biology, marine biology and anatomy &
physiology. This lab is an ideal choices for classroom studies of
mutagen and teratogen. Because zebrafish development
is very rapid, from zygote to multicellular free-swimming fish in
three to four days. Mutation are easily observed.
The embryos are extremely transparent. Students
can easily observe a heart beat, formation of eye, development of the nervous
system, skeletal system, muscle and circulatory system.
Glossary
Notes to the Teacher: to top
PREPARATION
Your are to provide dilution concentration of chemical (teratogen or
mutagen)
Three labs periods of approximately, 55 minutes
class periods to complete orientation (introduction)
day 1- lecture on the characteristic of zebrafish
- zebrafish embryology
day 2 - review the procedure student will be undertaking
day 3- removal, separation and observation of egg
- experiment
additional time is subsequent periods can be used for
discussion and review.
Care
Zebrafish are available at pet stores through out the nation. They
can be most easily maintained in 10 gallon aquarium at a temperature of
28C. Zebrafish will not breed at temperature above 31C or below 25C.
Keep fish in good breeding condition by feeding them dry flake food like
Tetra brand and live adult brine shrimp.
Numerous labs and information concerning the Zebrafish are available through the Internet and Medical/Research libraries.
"The Zebra Book"- A Guide for the Laboratory Use of
Zebrafish, Monte Westerfield,
University of Oregon Press, 1993
http://zfish.uregon.edu/zf_info/zfbook/zfbk.html
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GENERAL METHODS FOR ZEBRAFISH BREEDING to top
BREEDING
University
of Oregon Breeding information
BREEDING
SCHEDULE FOR MAXIMAL EMBRYO PRODUCTION
GENERAL
METHODS FOR ZEBRAFISH CARE 1
GENERAL
METHODS FOR ZEBRAFISH CARE 2
MICROSCOPIC
OBSERVATION (embryo development)
REMOVING EMBRYO FROM THE CHORION
EMBRYO DEVELOPMETN (Zygote Period)
Bellow are collection of brief description of all the stages in
the web pages and terms of important for staging:
http://zfish.uoregon.edu/zf_info/zfbook/stages/figs/fig1.html
http://zfish.uoregon.edu/zf_info/zfbook/stages/figs/fig39.html
http://zfish.uoregon.edu/zf_info/zfbook/stages/figs/fig39a.jpg
http://zfish.uoregon.edu/zf_info/zfbook/stages/figs/fig39b.jpg
Stage - h - HB - Description
Zygote period
1-cell - 0 - 1,2 - Cytoplasm streams towards animal pole to form the blastodisk
Cleavage period
2-cell - 3/4 - 3 - partial cleavage
4-cell - 1 - 4 - 2 x 2 array of blastomeres
8-cell - 1 1/4 - 5 - 2 x 4 array of blastomeres
16-cell - 1 1/2 - 6 - 4 x 4 array of blastomeres
32-cell - 1 3/4 - 7 - 2 regular tiers (horizontal rows) of blastomeres,
sometimes in 4 x 8 array
64-cell - 2 - 8 - 3 regular tiers of blastomeres
Blastula period
128-cell - 2 1/4 - 9 - 5 blastomere tiers; cleavage planes irregular
256-cell - 2 1/2 - - 7 blastomere tiers
512-cell - 2 3/4 - - 9 tiers of blastomeres; NO: YSL forms
1k-cell - 3 - 10 - 11 tiers of blastomeres; NO: single row of YSL nuclei;
slight blastodisk cell cycle asynchrony
High - 3 1/3 - - >11 tiers of blastomeres; beginning of blastodisk
flattening; NO: YSL nuclei in two rows; substantial division
asynchrony
Oblong - 3 2/3 - 11 - Flattening produces an elliptical shape; NO:
multiple rows of YSL nuclei
Sphere - 4 - 12 - Spherical shape; flat border between blastodisk and
yolk
Dome - 4 1/3 - 13 - Shape remains spherical; yolk cell bulging (doming)
towards animal pole as epiboly begins
30%-epiboly - 4 2/3 - 14 - Blastoderm an inverted cup of uniform thickness;
margin reaches 30% of distance between the
animal and vegetal poles
Gastrula period
50%-epiboly - 5 1/4 - - Blastoderm remains uniform in thickness Germ-ring
- 5 2/3 - - Germ ring visible from animal pole;
50%-epiboly
Shield - 6 - 15 - Embryonic shield visible from animal pole, 50%-epiboly
75%-epiboly - 8 - 16 - Dorsal side distinctly thicker; epiblast, hypoblast,
evacuation zone visible
90%-epiboly - 9 - - Brain rudiment thickened; notochord rudiment distinct
from segmental plate
Bud - 10 - 17 - Tail bud prominent; notochord rudiment distinct from
neural keel; early polster; midsagittal groove in anterior
neural keel; 100%-epiboly
Segmentation period
1-somite - 10 1/3 - - First somite furrow
5-somite - 11 2/3 - 18 - Polster prominent; optic vesicle, Kupffer's
vesicle
14-somite - 16 - 19 - EL = 0.9 mm; otic placode; brain neuromeres,
v-shaped trunk somites; YE barely forming; NO:
pronephric duct
20-somite - 19 - 20 - EL = 1.4 mm. YE/YB > 0.5 and < 1; muscular
twitches; lens, otic vesicle, rhombic flexure; hindbrain
neuromeres prominent; tail well extended
26-somite - 22 - - EL = 1.6 mm; HTA = 125 degrees; Side-to-side flexures;
otoliths; Prim-3
Pharyngula period
Prim-5 - 24 - - EL = 1.9 mm; HTA = 120 degrees; OVL = 5; YE/YB = 1;
early pigmentation in retina and skin; median fin
fold; red blood cells on yolk, heart beat
Prim-15 - 30 - - EL = 2.5 mm; HTA = 95 degrees; OVL = 3; YE/YB > 1;
YB/HD = 2; early touch reflex and reduced
spontaneous movements; retina pigmented; dorsal stripe to somite 12;
weak circulation; caudal artery halfway to end of tail;
caudal vein braided; shallow pectoral fin buds; straight tail; NO:
cellular degeneration at end of tail; circulation in aortic arch 1
Prim-25 - 36 - - EL = 2.7 mm; HTA = 75 degrees; OVL = 1; PF(H/W) =
3/4; early motility; tail pigmentation and ventral
stripe filling out; strong circulation; single aortic arch pair; caudal
artery is 3/4 of the way the to the end of tail; pericardium not
swollen; NO: PF apical ectodermal ridge
High-pec - 42 - - EL = 2.9 mm; HTA = 55 degrees; OVL < 1 and > 1/2;
YE/YB = 1.5; YB/HD < 1.3; PF(H/W) = 1;
dechorionated embryos rest on side after swimming; YE remaining cylindrical;
PF apical ridge prominent; early lateral stripe;
complete dorsal stripe; xanthophores in head only; iridophores in retina
only; pericardium prominent; HO: heart chambers;
segmental blood vessels; mandibular and hyoid arches; foregut developments
olfactory cilia; thickened otic vesicle walls
Hatching period
Long-pec - 48 - - EL = 3.1 mm; HTA = 45 degrees; OVL = 1/2; PF(H/W)
= 2; resting dorsal up; YE beginning to taper; PF
pointed; dorsal and ventral stripes meet at tail; ca. 6 melanophores
in lateral stripe; iridophores plentiful on retina; distinct yellow
cast to head; NO: circulation in 2-4 aortic arches and in segmental
vessels; olfactory cilia beating; semicircular canals;
neuromasts
Pec-fin - 60 - - EL = 3.3 mm; HTA = 35 degrees; movements too rapid
to resolve; YB tapering into YE; up to 10
melanophores in lateral stripe; PF flattenened into fin shape with
prominent circulation; iridophore retinal ring fills out;
iridophores in dorsal stripe; NO: PF cartilage and actinotrichia; gut
tract; 2 chambers in otic vesicle; early jaw cartilages;
circulation in 5-6 aortic arches; mouth remaining small and open at
ventral location midway between eyes
Protruding-mouth - 72 - - EL = 3.5 mm; HTA = 25 degrees; wide open
mouth protruding anterior to eye; iridophores in yolk
stripe; eye half covered by iridophores; dorsal body as yellow as head;
NO: gill slits and filament buds; cartilage in branchial
arch 1 and 5; operculum covers the branchial arch 1 or 2; cleithrum
Abbreviations: EL: embryo length, PF: pectoral fin, h: hours of development
at 28.5C. HB: approximate stage number in the
Hisoaka & Battle (1958) zebrafish staging series (reasonably accurate
through HB stage 20), HD: head diameter in dorsal
view, NO: Nomarski optics, H/W: height/width, Prim: Prim stages refer
to the number of the myotome to which the leading
end of the posterior lateral line primordium has advanced. YB: yolk
ball, YE: yolk extension, YSL: yolk syncytial layer
(These sketches have been scanned at low resolution. You can obtain
higher resolution sketches by clicking on the figure you
want. Each high resolution file is approxiately 100-200 kb in size,
so allow time for down-loading. The computer files are also
available for downloading by holding down the "option" key (Mac) or
"shift" key (PC) while clicking on the figure you want. If
you would like printed copies of these figures, contact C. Kimmel kimmel@uoneuro.uoregon.edu.)
1-cell: 2-cell: 4-cell: 8-cell: 16-cell:
32-cell: 64-cell: 128-cell: 256-cell: 512-cell:
1k-cell: high: oblong: sphere:
dome: 30%-epiboly: 50%-epiboly: germ ring:
shield: 8h: 9h: 10h: 11h:
12h: 14h: 16h: 18h: 19.5h:
22h: 25h: 31h: 35h: 42h:
48h: 60h: 72h:
http://zfish.uoregon.edu/zf_info/zfbook/stages/figs/fig39.html
http://zfish.uoregon.edu/zf_info/zfbook/stages/figs/fig39a.jpg
http://zfish.uoregon.edu/zf_info/zfbook/stages/figs/fig39b.jpg
Stage - h - HB - Description
animal pole: location on the egg where the polar bodies emerge, corresponding
to the point of
fertilization in fish like zebrafish, just below where the sperm penetrates
the chorion through the
micropile (passage way)
animal-vegetal axis: a line passing through the animal and vegetal poles
of the embryo before the
end of epiboly
anterior: towards the front (or head)
anterior horn: the distinctive anterior region of the ventral stripe
of melanophores, developing
between the ear and eye
anterior-posterior (AP) axis: the principal axis of the embryo, here
synonymous with rostrocaudal
axis and embryonic axis
aortic arch: artery leading from the ventral aorta to the paired radix
(root) of the dorsal aorta, or,
for the first two arches, to the internal carotid artery; an aortic
arch develops in all but the most
posterior of the seven pharyngeal arches, and the last four will carry
the blood supply to and from
the gills
AP: anterior-posterior
atrium: heart chamber collecting venous blood from the sinus venosus
and delivering it to the
ventricle; generates the first of each (doubled) heart beat
axial hypoblast: hypoblast that consists of mesodermal and probably
endodermal precursor cells
developing in the dorsal midline; includes prechordal plate and chorda
mesoderm
axis: a line, or alternatively shorthand for the anterior-posterior or embryonic axis
blastoderm: cellular part of the embryo, excluding the yolk cell, derived
from the blastodisc by early
morphogenesis; refers particularly to the time when the cell array
is sheet-like, between 30%-epiboly
and the end of gastrulation
blastodisc: (-disk) dome of cytoplasm (disk-like in the case of larger
teleost eggs such as Fundulus
and Salmo) that segregates from the yolk towards the animal pole during
and after the one
cell-stage, and which undergoes cleavage
blastomere: a cell arising during cleavage; the term encompasses the
partially cleaved, incomplete
"cells" at the blastodisc margin before they collectively form the
yolk syncytial layer in the midblastula
blastula: classically the single-layered hollow ball of cells formed
by cleavage in organisms that
show this developmental style; here used to mean a stereoblastula,
not hollowed-out, and as a
descriptor for the period of development when the blastodisc begins
to look ball-like, at the 128-cell
stage through the time of onset of gastrulation
blood island: nest of developing blood cells arising late in the segmentation
period from the
intermediate mass, and located in the anterior-ventral tail, just posterior
to the yolk extension
Brachet's cleft: the visible division between epiblast and hypoblast in the gastrula
branchial arch: gill arch; the last five of the set of seven pharyngeal
arches; the numbering system
can be confusing; generally branchial arch #1 is the first gill arch,
or the third pharyngeal arch, but
some authors do not follow this convention
cardinal vein: bilaterally paired longitudinial vein; the anterior cardinal
returns blood from the head,
and the posterior cardinal returns it from the trunk; these two vessels
join together on each side as
the common cardinal vein (duct of Cuvier; misnamed the vitelline vein)
that leads across the yolk cell
to the heart's sinus venosus
carotid artery: see internal carotid artery
caudal: pertaining to the tail, or the posterior direction
caudal artery: extension of the dorsal aorta in the tail
caudal vein: vein in the tail returning blood from the trunk and tail
to the heart, leads directly into the
axial vein in the posterior trunk
central canal: fluid-filled narrow cavity in the spinal cord
central nervous system: the brain and spinal cord
cerebellum: specialized brain region derived from the dorsal metencephalon
(anterior hindbrain, and
perhaps including posterior midbrain) and becoming distinctive late
in the segmentation period
chorion: the egg shell
cleavage: an early mitotic cell division occurring in the blastodisc,
special in that the cell cycles are
short in length, are not accompanied by cell growth during interphase,
and occur synchronously or
metasynchronously with other cleavages of the same number; in the staging
series, the cleavage
period refers to the period of development encompassing the first 6
six zygotic cell cycles
coelom (coelomic cavity): fluid-filled mesodermally lined cavity separating
visceral organs including
the heart from the body wall
common cardinal vein: see cardinal vein
convergence: deep cell movement toward the dorsal side of the embryo
during the gastrula and
early segmentation periods
deep cell, a cell in the the blastodisc (first at the 64-cell stage)
or blastoderm that is completely
covered over by other cells, the outermost being cells of the enveloping
layer
diencephalon: the more posterior and ventral of two forebrain neuromeres,
the other being the
telencephalon; major derivatives are the eye cups, the brain pretectal
region, the thalamus,
hypothalamus, and epithalamus (including the habenula and epiphysis)
dorsal: toward the back (the side opposite to the belly)
dorsal aorta: principal unpaired, median artery of the trunk, leading
from the paired roots (radices)
of the dorsal aorta to the caudal artery
dorsal-ventral (dorsoventral): axis passing from the back to the belly;
within a sagittal plane and at
right angles to the anterior-posterior axis
embryo length (EL): at any stage the embryo's longest linear dimension
embryonic axis: see anterior-posterior axis
endothelium: epithelial lining of any blood vessel including the heart
enveloping layer (EVL): outermost monolayer of cells surrounding the
embryo that become very
flattened in the blastula and give rise to the periderm
epiblast: the outer of the two layers of the blastoderm that form during
gastrulation, corresponding
to primitive ectoderm during gastrulation and to the definitive ectoderm
after gastrulation
epiboly: the thinning and spreading of both the YSL and the blastoderm
over and across the yolk
cell, eventually encompassing the yolk cell completely; epiboly begins
at dome stage, converts the
blastodisc to the blastoderm, and is considered to be over when the
yolk plug closes over (at
100%-epiboly)
epithelium: a compact and sheet-like arrangement of cells, polarized
with the apical surface to one
side (primitively the outside) and the basal surface to the other
EVL: enveloping layer
external yolk syncytial layer (E-YSL): portion of the YSL that is outside
of the blastoderm
margin during epiboly
forebrain: the most anterior region the brain including both the telencephalon
and diencephalon; we
have not observed an early transient stage when the forebrain is distinguished
from the midbrain but
has not subdivided, that would correspond to the prosencephalon in
tetrapods
gastrula: classically a postblastula stage in which an archenteron (primitive
gut or gastrocoele) forms
by invagination or involution of cells through a blastopore and when
the germ layers and embryonic
axis appear; the zebrafish forms neither an archenteron nor a blastopore,
and here the term refers to
the roughly equivalent period of development, beginning at the onset
of involution (at the
50%-epiboly stage) that produces the two primary germ layers, the epiblast
and hypoblast, and
during which the definitive embryonic axis forms by convergence and
extension movements
gastrulation: morphogenesis during the gastrula period
gill arch: one of the subset of pharyngeal arches (pharyngeal arches
3-6, or branchial arches 1-4)
that will develop gills
gill filament: branched region of the gill where respiratory exchange takes place
hair cell: specialized neuronal receptor cell of the lateral line and acoustico-vestibular systems
hindbrain: the most posterior of the three principle regions of the
brain, forming the
rhombencephalon and all or most of the metencephalon
horizontal: during cleavage and blastula periods a plane perpendicular
to the animal-vegetal axis;
later a longitudinal plane parallel to the embryonic axis and perpendicular
to the dorsal ventral axis,
i.e. at right angles to both transverse and sagittal planes
horizontal myoseptum: a connective tissue partition developing at the
apex of the chevron-shaped
myotome and separating dorsal (epaxial) and ventral (hypaxial) body
wall muscle masses
hypoblast (mesendoderm): the inner of the two layers of the blastoderm
that forms during
gastrulation and give rise to the definitive mesoderm and endoderm
hypothalamus: a specialized brain region of the ventral diencephalon
arising near the end of the
segmentation period; the embryonic hypothalamic region will give rise
to the posterior pituitary gland
as well as a number of brain nuclei
intermediate mass: the very early blood rudiment located deep to the
somites in the posterior trunk
at a stage before the blood cells collect into the (more prominent)
blood island
internal carotid artery: artery originating at the junction of the first
two aortic arches and supplying
the anterior brain
internal yolk syncytial layer (I-YSL): the portion of the YSL that lies
deep to the blastoderm
during epiboly
involution: deep cell movement at the blastoderm margin in which the
DEL folds inwards and back
upon itself, producing the germ ring and its two primary germ layers,
the epiblast and hypoblast
iridophore: reflective pigment cell
mandibular arch: first (most anterior) pharyngeal arch, forming the
principal elements of the jaw of
the early larva
Meckel's (mandibular) cartilage: ventral cartilage of the mandibular
arch forming the principal
support of the (lower) jaw
medial: toward the midline
median: at the midline
melanophore (melanocyte): a neural crest-derived cell containing black melanin pigment
mesenchyme: a mesh-like cell arrangement, less compact than an epithelium
divide less synchronously, and motility and zygotic transcription are first observed
midbrain (mesencephalon): the brain region between the forebrain anteriorly
and the hindbrain
posteriorly, including the tectum dorsally and the midbrain tegmentum
ventrally
midsagittal plane: the plane of bilateral symmetry, located at the midline
myotome: portion of the somite giving rise to body wall muscle masses
neural crest: a cell population arising from the dorsolateral aspect
of the central nervous system
primordium during the segmentation period, and later migrating along
stereotyped pathways to give
rise to a diverse and well-defined set of cell types including pigment
cells, peripheral neurons and
glia, and head cartilage
neural groove: a midsagittal depression on the surface of the anterior
neural plate present during the
early segmentation period
neural plate: the earliest recognizeable dorsal ectodermal primordium
of the central nervous system
present near the end of gastrulation before infolding to form the neural
keel; consists of a thickened
pseudostratified epithelium
neural rod: an intermediate stage in the development of the central
nervous system present during
the segmentation period; the neural rod is roughly cylindrical in shape,
forms from the neural keel,
and is not yet hollowed out into the neural tube
neural tube: cavity-containing primordium of the central nervous system,
developing from the neural
rod in the late segmentation period
notochord: rod-like principal supportive element of the embryo and larva,
present in the midline just
ventral to the neural tube, and differentiating during the segmentation
period to form large vacuolated
principal cells and a surrounding thin epithelial notochord sheath
optic primordium: lateral outgrowth from the forebrain that will form
the eyeball (excluding the
lens); equivalent to the optic vesicle of tetrapods, but apparently
not a hollow structure; develops
into the two layered optic cup
optic tectum: the roof of the midbrain, morphologically visible by the end of the segmentation period
paraxial hypoblast: hypoblast that is mainly or entirely mesodermal,
positioned laterally to the axial
hypoblast; forms somites and their derivatives in the trunk and muscles
and endothelium in the head
pericardium: portion of the coelomic cavity present as a distinctive chamber surrounding the heart
peripheral nervous system: nervous structures including ganglia outside
of the central nervous
system
pharyngeal (visceral) arch: a segment of the lateral wall of the pharynx
that will form jaw
structures (anterior two arches ) or gill structures (posterior 5 arches);
an arch includes a compact
mesenchyme lined by inner endoderm and outer epidermis; each arch is
separated from neighboring
arches by an endodermal outpocketing (a pharyngeal pouch) meeting a
slight ectodermal inpocketing
(a pharyngeal cleft) where a gill slit develops during the hatching
period (except between the first and
second arches)
pharyngula: generally, a vertebrate embryo that has developed to the
phylotypic stage; in the series
used as a period name to describe the second of the three days of embryonic
development
pharynx: swollen region of the anterior foregut, posterior to the mouth
and anterior to the liver; its
walls form the jaws and gills
phylotypic stage: the stage at which the embryo develops features defining
it as a vertebrate or
chordate, including the notochord, neural tube, pharyngeal arches,
somites and postanal tail
retina: the portion of the eye developing from the optic primordium
and including the neural retina
and the retinal pigment layer
rostral: toward the head, for the zebrafish embryo synonymous with anterior
rostral-caudal (rostrocaudal) axis: here synonymous with anterior-posterior axis
sagittal: a plane parallel to the plane of bilateral symmetry
sclerotome: medial ventral region of the somite that will form vertebral cartilages
segmental artery: artery leading from the dorsal aorta or caudal artery
to the spinal cord; the
arteries alternate in adjacent segments with segmental veins
segmental vein: vein leading from the spinal cord to the caudal vein,
axial vein, or posterior cardinal
vein
segmentation: a repetition of elements, particularly along the AP axis;
used in the series to define
the period of development between the gastrula and pharyngula
sinus venosus: heart region collecting blood from the paired common
cardinal veins and delivering
to the atrium
somite: undifferentiated mesodermal component of an early trunk or tail
segment or metamere,
derived from paraxial mesoderm; forms the myotome, sclerotome and perhaps
dermatome
subclavian artery: artery supplying the pectoral fin
subclavian vein: vein returning blood from the pectoral fin to the cardinal system
telencephalon: the anterior and dorsal forebrain neuromere, includes the olfactory bulb
vegetal pole: location on the egg opposite to the animal pole, corresponding
later to the point on the
yolk cell furthest from the developing blastodisc
ventral: toward the belly (or yolk)
ventral aorta: outflow artery from the heart to the aortic arches
ventricle: fluid-filled brain cavity; alternatively a heart chamber
collecting venous blood from the
atrium and delivering it to the conus arteriosus; the ventricle mediates
the second and major
component of each (doubled) heart beat
vertical: here meaning a plane parallel to the animal-vegetal axis during
the cleavage and blastula
periods
visceral: pertaining to the gut or endoderm or splanchnic mesoderm associated with endoderm
yolk: nutrient store for embryonic development in the form of semicrystalline
phospholipoprotein and
contained within yolk granules
yolk ball: the anterior round region of the yolk cell present after
the yolk extension forms during the
segmentation period
yolk extension: the posterior elongated region of the yolk cell that
forms during the segmentation
period
yolk cell: giant syncytial uncleaved cell containing the yolk; underlies
the blastodisc early, and
becomes enveloped by the blastoderm during epiboly
yolk granule: membrane-bounded sac, of the order of 50 µm in diameter,
containing yolk; yolk
granules are packed densely in the interior of the yolk cell, deep
to its syncytial layer, and make up
the great bulk of its total volume
yolk plug: the bit of yolk cell protruding beyond the blastoderm margin
in the late gastrula before
epiboly is complete
yolk stripe: a late-forming melanophore stripe along the median ventral
aspect of the yolk ball and
particularly the yolk extension
yolk syncytial layer (YSL): peripheral layer of the yolk cell including
nuclei and non-yolky
cytoplasm
YSL: yolk syncytial layer
zygote: the fertilized egg, defining in the series the period between
fertilization and the end of the first
clea
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THE SCIENTIFIC METHOD
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Title
The project title should give information regarding the topic being
studied. It may consist of the actual problem statement.
Purpose
The purpose of this activity is to demonstrate basic research.
In this investigation student will view morphological changes induce by
chemical into developing Zebrafish eggs.
Hypothesis
After getting information about the investigation, student(s) should
make an educated guess about what they think the answer to the question
may be.
example: The hypothesis of this investigation is that...
Material
There should be a list of all the materials that are used, preferably
in column form. Amount of materials should always be indicated in metric
units.
Procedure
The Procedures should be listed step by step, amounts involved
should be included
Manipulated variable
is the item that is intentionally changed in order to test it.
Responding variable
is what is changed in response to the manipulated variable.
Constant variable
are all other factors in the investigation that must remain the same.
Data
All information that is collected include graphs, chart, or pictures.
Each step of this investigation should be documented as to the date, time,
place, students who participated.
Results
The investigator(s) should state the finding of the investigation based
on the data that has been observed and carefully analyzed.
example: According to the data...
Conclusion
A statement should be made on whether or not the results support
the hypothesis. The investigator(s) should discuss how specific data
from the experiment supported the hypothesis and describe problems
that might have affected the results.
Applications
The student(s) should explain why this experiment was important (relevant
to real life situations).
| Student lab
|
|
|
| blastodisc | gastrula |
| blastula | gill arch |
| chorion | holoblastic cleavage |
| cleavage | hypoblast |
| coelon | meroblastic cleavage |
| differentiation | median |
| dorsal | mesenchyme |
| endothelium | notochord |
| epiblast | somite |
| epibody | yolk |
| fertilization |
In this investigation you will:
1. Identify the parts, functions and, proper care, of the equipments.
2. Identify appropriate safety procedures use
with mutagen and teratogen.
3. Use careful observations and exploratory activities
to identify variables and develop problem
statements.
4. Compare mitosis and meiosis with regard to chromosome
number in parent cells versus
daughter cells, types of cells
produced, total number of cells produced, and the
number of divisions.
5. Identify and differentiate the major characteristics of vertebrate development.
6. Explain the relationships among the number of eggs,
methods of fertilization, and rates of
embryonic development as related
to species survival.
7. After completing this investigation, you are to write a " Research
Paper" (include the following
items: purpose, hypothesis, material,
procedure, variable, data, results, conclusion and
application).
| Materials & Equipment Needs |
| MICRODISSECTION INSTRUMENTS. | |||
| Aquarium | Videomicroscope | Chemically treated water | Forceps |
| Aquarium hood with light | Nicotine | Metric ruler | Scalpels |
| Acetic acid | Estrogen | Micropipet 10uL | Scissors |
| Aquarium heater | 0.85% Ethanol | Zebrafish | |
| Aquatic net | 1.00% Ethanol | Zebrafish eggs | |
| Culture dish | 2.00% Ethanol | Brine shrimp | |
| Marbles | 2.85% Ethanol | Glass pipettes | |
| Siphon hose | Slides | ||
| Compound Microscope | Microscope depression slides | ||
| Stereomicroscope |
STUDENT PREPARATION
| ALTERNATIVE ALCOHOL PROCEDURE |
I. Observation and Measuring Eggs
1. Student(s) observe draw, and measure development
of eggs over a period of a week.
2. Student(s) observed and draw, how embryo feed themselves.
3. Place egg in depression slide with egg's solution
and observed using compound microscope.
4. Describe the formation of organs structure as they
develop.
5. Graph the embryo length(mm) versus time(hr).
Graph the embryo heart rate versus length(mm).
Graph
the embryo heart rate versus time(min).
II. OBSERVATION OF EMBRYOLOGICAL DEVELOPMENT
1. Identify the neural tube, brain , somite,
heart and eyes of a developing embryo.
(label these on all your
drawing if present)
2. Compare and contrast between cleavage of
the control group versus experimental.
3. What difference do you see between cleavage of
the control group vs experimental?
4. Compare and contrast Cleavage, Blastula, Gastrula
and Hatching between the control
versus experimental.
5. How would you enhance this investigation?
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Corley-Smith,GE,Lim,C.J.,& Branhorst,B.P., "The Zebrafish,"
Institute of Molecular Biology and Biochemistry Science Monitor, Vol. 3(5)
Cronkite, Donald, Professor of Biology, Hope College
cronkite@hope.edu.
Mac Vicar,J. : Antenatal detection of fetal abnormality;
Physical Methods,
Br. Med. J. 32:4,,1976
Mc Glone, Barbara: Zebra, Zebra, Where Are Your Stripes?
Living Computer?
Woodrow Wilson National Fellowship, Institute 1995
Mc Keown,T.: Human Malformations: Introduction. Br. Med. J. 32:1,1976
Moore, Keith. The Developing Human Clinical Oriented Embryology
W.B. Saunders, 1982.
Oppenheimer, Steven B. Embryological Research, The Science
Teacher, Vol. 56,pp40-43,Nov.1989
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