Cryopreservation of reproductive products of many aquatic species has been successfully achieved. ... Cryopreservation technology applied to the preservation of fish gametes in aquaculture plays an important role in seed production, genetic management of broodstock and conservation of aquatic resources
Cryopreservation and its application to aquaculture.pptxNarsingh Kashyap
What is Cryopreservation ?
Cryopreservation is a process where biological materials such as cells and tissues are preserved by cooling to very low temperatures, usually at -196°C (the temperature of liquid nitrogen), yet remain viable after later warming to temperatures above 0°C.
Cryopreservation in aquatic species goes back 65 years and began about the same time as similar research was performed in livestock (Blaxter 2011).
In India, NBFGR & CIFA are the primary organization carrying out fish sperm cryopreservation for long term gene banking (J. K. Jena 2012)
Fishes, amphibians, reptiles, and birds have paired pharyngeal ultimobranchial glands that secrete the hypocalcemic hormone calcitonin. The corpuscles of Stannius, unique glandular islets found only in the kidneys of bony fishes, secrete a peptide called hypocalcin.
Carps form the mainstay of aquaculture in India contributing over 85% of the total aquaculture production There are 61,259 species of vertebrates recognized world; over 30,700 are fish species of which 8,411 ore fresh water while 11,650 are marine. In India 2,163 spp. are fin fishes have been recorded from upland cold water (157; 7.26%) warm water of the plain (54; 20.99%), Brackish water (182; 8.41%) and marine environment (1,370; 63.43%). Some of these species are cultured at commercial level which covering a lot varieties of fin fishes The three Indian major carps, namely Catla (Catla catla), Rohu (Labeo rohita) and Mrigal (Cirrhinus mrigal) contribute the bulk of production to the extent of 75 to85 percent of the total fresh water fish production, the three exotic carp such as Silver carp (Hypophthalmichthys molitrix) and Common carp (Cyprinus carpio ), Grass carp (Ctenopharyngodon idella )form the second important group to incorporate several other medium and minor carp and into the carp poly culture system several method were used because of their region specific consumer preference and higher market demand.
History
Carp culture in India was restricted only to a homestead backyard pond activity in west Bengal and Odisha until late 1950 s with seed from riverine sources as the only input resulting low level of production the technological breakthrough breeding of carp through hypophysation in 1957 freshwater aquaculture of the country the country till 1984 virtually laid the foundation of scientific carp farming in the country.
Important characteristics of Indian major carps:-
Indian major carp grow fast and can reproduce even in artificial ponds. They feed upon phytoplankton, zooplankton, decaying organic matter, aquatic plant etc. stomach is absent in the alimentary canal of the major carps. Three types of Indian major carps are cultured in
A fishing gear is the tool with which aquatic resources are captured, whereas the fishing method is how the gear is used. Gear also includes harvesting organisms.
Dr. K. Rama Rao
Department of Zoology
Govt, Degree College
TEKKALI; Srikakulam Dt: A.P.
the presentation provides details regarding the natural and artificial feeds of fishes, purified and semipurified diets, feeds based on the moisture contents, the larval feeds including the most recent spray dried and vacuumdried feeds, microparticulate diets, the microencapsulated, the microcoated and the microbound diets, microextruded marumerisation, and particle associated rotated agglomeration
1975; Scott and Baynes, 1980; Chao et al., 1987; Baynes and Scott, 1987; Koldras and Bienarz, 1987; Harvey and Kelley, 1988; Leung and Jamieson, 1991; Gwo, et al., 1993; Rana 1995; Babiak et al., 1997; Akcay et al., 2004). Extenders and cryoprotectants are important and play a vital role in cryopreservation. Irrespective of the species, fish semen requires dilution before it has to be cryopreserved. Extenders used for diluting the fish semen are generally designed to be compatible with the physico-chemical composition of seminal fluid of the candidate species. The chemical constituents of extenders vary enormously (Scott and Baynes, 1980; Stoss, 1983). A range of cryoprotective agents of permeating and non-permeating categories are available for the use to minimize cryoinjuries during cooling and thawing process. The DMSO and glycerol are widely used cryoprotective agents. Suitability of extenders and cryoprotectants differs from one fish to another (Muchlisin, 2005). Semen is commonly packaged in cryovials (Ott and Horton, 1971), plastic straws (Erdhal, 1986; Chao et al., 1987) or visotubes (Mounib, 1978; Stein and Bayrle, 1978) cooled over liquid nitrogen vapor or in programmable freezer and stored in liquid nitrogen (Cognet, et al., 1996). Fish semen can also be cryopreserved as pellets on dry-ice blocks and then stored in caped cryovials in liquid nitrogen (Leung and Jamieson, 1991). Various cooling methods have been successfully used to cryopreserve the fish sperm. Careful manipulation of temperature excursion is required to control the size, configuration and location of ice crystals. Thus choice and concentration of cryoprotectants and rate of cooling is needed to be optimized for each species as the basis for any protocol development.
From the current state of art of fish spermatozoa cryopreservation and species differences, one universal protocol cannot be suggested since response to cryoprotectant and freezing vary with the different biology. Thus, optimization of the protocol is needed for each individual species though some general rules are applied for each fish species. In the present communication, basic principles and essential steps of cryopreservation techniques for the sperm of fresh water fish species are explained with the example from a snowtrout species
(S.richardsonii) as a model. For the development of any reliable protocols for fish semen cryopreservation, emphasis should always be placed on the standardization.
CRYOBIOLOGIC PRINCIPLES
Nature dictates that biological material will decay and die. The structure and function of organisms are changed and lost with the time. An attempt to stop the biological clock, experiments with temperature and water contents of the cell is the basic theme of cryopreservation research. The use of much lower temperatures has proved a means of storing living organisms in a state of suspended animation for extended periods. The removal of water from biological material in the frozen state
Cryopreservation and its application to aquaculture.pptxNarsingh Kashyap
What is Cryopreservation ?
Cryopreservation is a process where biological materials such as cells and tissues are preserved by cooling to very low temperatures, usually at -196°C (the temperature of liquid nitrogen), yet remain viable after later warming to temperatures above 0°C.
Cryopreservation in aquatic species goes back 65 years and began about the same time as similar research was performed in livestock (Blaxter 2011).
In India, NBFGR & CIFA are the primary organization carrying out fish sperm cryopreservation for long term gene banking (J. K. Jena 2012)
Fishes, amphibians, reptiles, and birds have paired pharyngeal ultimobranchial glands that secrete the hypocalcemic hormone calcitonin. The corpuscles of Stannius, unique glandular islets found only in the kidneys of bony fishes, secrete a peptide called hypocalcin.
Carps form the mainstay of aquaculture in India contributing over 85% of the total aquaculture production There are 61,259 species of vertebrates recognized world; over 30,700 are fish species of which 8,411 ore fresh water while 11,650 are marine. In India 2,163 spp. are fin fishes have been recorded from upland cold water (157; 7.26%) warm water of the plain (54; 20.99%), Brackish water (182; 8.41%) and marine environment (1,370; 63.43%). Some of these species are cultured at commercial level which covering a lot varieties of fin fishes The three Indian major carps, namely Catla (Catla catla), Rohu (Labeo rohita) and Mrigal (Cirrhinus mrigal) contribute the bulk of production to the extent of 75 to85 percent of the total fresh water fish production, the three exotic carp such as Silver carp (Hypophthalmichthys molitrix) and Common carp (Cyprinus carpio ), Grass carp (Ctenopharyngodon idella )form the second important group to incorporate several other medium and minor carp and into the carp poly culture system several method were used because of their region specific consumer preference and higher market demand.
History
Carp culture in India was restricted only to a homestead backyard pond activity in west Bengal and Odisha until late 1950 s with seed from riverine sources as the only input resulting low level of production the technological breakthrough breeding of carp through hypophysation in 1957 freshwater aquaculture of the country the country till 1984 virtually laid the foundation of scientific carp farming in the country.
Important characteristics of Indian major carps:-
Indian major carp grow fast and can reproduce even in artificial ponds. They feed upon phytoplankton, zooplankton, decaying organic matter, aquatic plant etc. stomach is absent in the alimentary canal of the major carps. Three types of Indian major carps are cultured in
A fishing gear is the tool with which aquatic resources are captured, whereas the fishing method is how the gear is used. Gear also includes harvesting organisms.
Dr. K. Rama Rao
Department of Zoology
Govt, Degree College
TEKKALI; Srikakulam Dt: A.P.
the presentation provides details regarding the natural and artificial feeds of fishes, purified and semipurified diets, feeds based on the moisture contents, the larval feeds including the most recent spray dried and vacuumdried feeds, microparticulate diets, the microencapsulated, the microcoated and the microbound diets, microextruded marumerisation, and particle associated rotated agglomeration
1975; Scott and Baynes, 1980; Chao et al., 1987; Baynes and Scott, 1987; Koldras and Bienarz, 1987; Harvey and Kelley, 1988; Leung and Jamieson, 1991; Gwo, et al., 1993; Rana 1995; Babiak et al., 1997; Akcay et al., 2004). Extenders and cryoprotectants are important and play a vital role in cryopreservation. Irrespective of the species, fish semen requires dilution before it has to be cryopreserved. Extenders used for diluting the fish semen are generally designed to be compatible with the physico-chemical composition of seminal fluid of the candidate species. The chemical constituents of extenders vary enormously (Scott and Baynes, 1980; Stoss, 1983). A range of cryoprotective agents of permeating and non-permeating categories are available for the use to minimize cryoinjuries during cooling and thawing process. The DMSO and glycerol are widely used cryoprotective agents. Suitability of extenders and cryoprotectants differs from one fish to another (Muchlisin, 2005). Semen is commonly packaged in cryovials (Ott and Horton, 1971), plastic straws (Erdhal, 1986; Chao et al., 1987) or visotubes (Mounib, 1978; Stein and Bayrle, 1978) cooled over liquid nitrogen vapor or in programmable freezer and stored in liquid nitrogen (Cognet, et al., 1996). Fish semen can also be cryopreserved as pellets on dry-ice blocks and then stored in caped cryovials in liquid nitrogen (Leung and Jamieson, 1991). Various cooling methods have been successfully used to cryopreserve the fish sperm. Careful manipulation of temperature excursion is required to control the size, configuration and location of ice crystals. Thus choice and concentration of cryoprotectants and rate of cooling is needed to be optimized for each species as the basis for any protocol development.
From the current state of art of fish spermatozoa cryopreservation and species differences, one universal protocol cannot be suggested since response to cryoprotectant and freezing vary with the different biology. Thus, optimization of the protocol is needed for each individual species though some general rules are applied for each fish species. In the present communication, basic principles and essential steps of cryopreservation techniques for the sperm of fresh water fish species are explained with the example from a snowtrout species
(S.richardsonii) as a model. For the development of any reliable protocols for fish semen cryopreservation, emphasis should always be placed on the standardization.
CRYOBIOLOGIC PRINCIPLES
Nature dictates that biological material will decay and die. The structure and function of organisms are changed and lost with the time. An attempt to stop the biological clock, experiments with temperature and water contents of the cell is the basic theme of cryopreservation research. The use of much lower temperatures has proved a means of storing living organisms in a state of suspended animation for extended periods. The removal of water from biological material in the frozen state
Know About Cryogenic Preservations And Its ApplicationsSpi-Mfg
Cryogenic preservation is storing and freezing embryos and is considered a crucial part of fertilization programs. So, if you're looking to buy the best cryogenic preservation products, make sure you get them from some reputable brands and platforms.
Similar to cryopreservation of fish gametes NBFGR gene bank (20)
Marine fisheries regulation act (mfra), and its amendments of gujrat and dama...Ashish sahu
Marine Fisheries Regulation Act
An Act to provide for the regulation of fishing by fishing vessels in the sea along the coast line of the State. (1) This Act may be called the Kerala Marine Fishing Regulation Act, 1980. (2) It extends to the whole of the State of Kerala.
Ocean acidification is the ongoing decrease in the pH value of the Earth's oceans, caused by the uptake of carbon dioxide (CO 2) from the atmosphere. ... When carbon dioxide dissolves into seawater, it forms carbonic acid (H2CO3). Ocean acidification impacts many species, especially organisms like oysters and corals.
Ocean acidification reduces the amount of carbonate, a key building block in seawater. This makes it more difficult for marine organisms, such as coral and some plankton, to form their shells and skeletons, and existing shells may begin to dissolve. ... The impacts of ocean acidification are not uniform across all species.
Mangroves and their importance with Threats iucn pptAshish sahu
A mangrove is a shrub or tree that grows in coastal saline or brackish water. ... Mangroves are salt-tolerant trees, also called halophytes, and are adapted to live in harsh coastal conditions. They contain a complex salt filtration system and a complex root system to cope with saltwater immersion and wave action.
Mangroves are defined as assemblages of salt tolerant trees and shrubs that grow in the intertidal regions of the tropical and subtropical coastlines. They grow luxuriantly in the places where freshwater mixes with seawater and where sediment is composed of accumulated deposits of mud.
Fisheries crime and their sollution pptAshish sahu
What is fisheries crime?: Fisheries crime is an ill-defined legal concept referring to a range of illegal activities in the fisheries sector. These activities – frequently transnational and organized in nature – include illegal fishing, document fraud, drug trafficking, and money laundering.
Illegal fishing is a key driver of global overfishing, it threatens marine ecosystems, puts food security and regional stability at risk, and is linked to major human rights violations and even organized crime.
Iconic species are socially, culturally and economically important, and the community expects them to be effectively managed and protected.
What is a flagship species? A flagship species is a species selected to act as an ambassador, icon or symbol for a defined habitat, issue, campaign or environmental cause. ... Flagship species are usually relatively large, and considered to be 'charismatic' in western cultures.
Collection device of fish eggs and larvae pdfAshish sahu
First, an incision is made in her abdomen and then a small tube is inserted. The farmer then puts his mouth on the tube and sucks out a sWhen the internal ovaries or egg masses of fish and certain marine animals are ripe for spawning they are called roe.mall quantity of eggs for examination. If they're the right color and consistency, he'll kill the animal and harvest the caviar.When the internal ovaries or egg masses of fish and certain marine animals are ripe for spawning they are called roe. I think the best way is to use a toothpick and place them on something flat like a small chunk of marble or any flat ornament. Then place in a submerged container. Don't let the eggs hit open air. Well, the eggs in my breeding tank hatched, but overnight the fry either escaped or died
Mollusca of India and need for conservationAshish sahu
Mollusca is the second-largest phylum of invertebrate animals after the Arthropoda. The members are known as molluscs or mollusks. Around 85,000 extant species of molluscs are recognized. The number of fossil species is estimated between 60,000 and 100,000 additional species
Goodbye, Gillnets Belize bans deadly fishing gear, protecting countless marine animals Magazine Opting Out Amazon packages are awash in plastic, and shoppers want alternatives Q&A with Sam Waterston On cod, climate change, and his ... Ocean Council Susan Rockefeller, Founder Kelly Hallman, Vice Chair Dede McMahon, Vice Chair Anonymous Samantha Bass Violaine and John Bernbach Rick Burnes Vin Cipol
A coral reef is an underwater ecosystem characterized by reef-building corals. Reefs are formed of colonies of coral polyps held together by calcium carbonate. Most coral reefs are built from stony corals, whose polyps cluster in groups.
Acharya Narendra Deva University of Agriculture and Technology is a university located in Kumarganj, Faizabad, Uttar Pradesh, India, established in 1975. It is named after the politician and educator Narendra Deva, who served as vice chancellor of the University of Lucknow and Banaras Hindu University. It also has constituent colleges in Ambedkar Nagar district and Azamgarh district, as well as a planned college in Gonda district.
Acharya Narendra Deva University of Agriculture and TechnologyAshish sahu
Acharya Narendra Deva University of Agriculture and Technology
Acharya Narendra Deva University of Agriculture and Technology is a university located in Kumarganj, Faizabad, Uttar Pradesh, India, established in 1975. It is named after the politician and educator Narendra Deva, who served as vice chancellor of the University of Lucknow and Banaras Hindu University. It also has constituent colleges in Ambedkar Nagar district and Azamgarh district, as well as a planned college in Gonda district.
Impact of climate change in atmosphere of oceanAshish sahu
How does climate change effect the ocean?
5 ways that climate change affects the ocean
Higher temperatures are bad for fish — and for us.
Polar ice is melting.
Rising sea levels represent a slow, seemingly unstoppable threat.
Warming oceans alter currents.
Climate change is affecting the chemistry of seawater.
Three main methods for pond management are adding aeration, water treatments like beneficial bacteria, and weed and algae controls. The single best water management strategy you can implement is adding pond aeration.
How to Create Map Views in the Odoo 17 ERPCeline George
The map views are useful for providing a geographical representation of data. They allow users to visualize and analyze the data in a more intuitive manner.
Operation “Blue Star” is the only event in the history of Independent India where the state went into war with its own people. Even after about 40 years it is not clear if it was culmination of states anger over people of the region, a political game of power or start of dictatorial chapter in the democratic setup.
The people of Punjab felt alienated from main stream due to denial of their just demands during a long democratic struggle since independence. As it happen all over the word, it led to militant struggle with great loss of lives of military, police and civilian personnel. Killing of Indira Gandhi and massacre of innocent Sikhs in Delhi and other India cities was also associated with this movement.
The Roman Empire A Historical Colossus.pdfkaushalkr1407
The Roman Empire, a vast and enduring power, stands as one of history's most remarkable civilizations, leaving an indelible imprint on the world. It emerged from the Roman Republic, transitioning into an imperial powerhouse under the leadership of Augustus Caesar in 27 BCE. This transformation marked the beginning of an era defined by unprecedented territorial expansion, architectural marvels, and profound cultural influence.
The empire's roots lie in the city of Rome, founded, according to legend, by Romulus in 753 BCE. Over centuries, Rome evolved from a small settlement to a formidable republic, characterized by a complex political system with elected officials and checks on power. However, internal strife, class conflicts, and military ambitions paved the way for the end of the Republic. Julius Caesar’s dictatorship and subsequent assassination in 44 BCE created a power vacuum, leading to a civil war. Octavian, later Augustus, emerged victorious, heralding the Roman Empire’s birth.
Under Augustus, the empire experienced the Pax Romana, a 200-year period of relative peace and stability. Augustus reformed the military, established efficient administrative systems, and initiated grand construction projects. The empire's borders expanded, encompassing territories from Britain to Egypt and from Spain to the Euphrates. Roman legions, renowned for their discipline and engineering prowess, secured and maintained these vast territories, building roads, fortifications, and cities that facilitated control and integration.
The Roman Empire’s society was hierarchical, with a rigid class system. At the top were the patricians, wealthy elites who held significant political power. Below them were the plebeians, free citizens with limited political influence, and the vast numbers of slaves who formed the backbone of the economy. The family unit was central, governed by the paterfamilias, the male head who held absolute authority.
Culturally, the Romans were eclectic, absorbing and adapting elements from the civilizations they encountered, particularly the Greeks. Roman art, literature, and philosophy reflected this synthesis, creating a rich cultural tapestry. Latin, the Roman language, became the lingua franca of the Western world, influencing numerous modern languages.
Roman architecture and engineering achievements were monumental. They perfected the arch, vault, and dome, constructing enduring structures like the Colosseum, Pantheon, and aqueducts. These engineering marvels not only showcased Roman ingenuity but also served practical purposes, from public entertainment to water supply.
2024.06.01 Introducing a competency framework for languag learning materials ...Sandy Millin
http://sandymillin.wordpress.com/iateflwebinar2024
Published classroom materials form the basis of syllabuses, drive teacher professional development, and have a potentially huge influence on learners, teachers and education systems. All teachers also create their own materials, whether a few sentences on a blackboard, a highly-structured fully-realised online course, or anything in between. Despite this, the knowledge and skills needed to create effective language learning materials are rarely part of teacher training, and are mostly learnt by trial and error.
Knowledge and skills frameworks, generally called competency frameworks, for ELT teachers, trainers and managers have existed for a few years now. However, until I created one for my MA dissertation, there wasn’t one drawing together what we need to know and do to be able to effectively produce language learning materials.
This webinar will introduce you to my framework, highlighting the key competencies I identified from my research. It will also show how anybody involved in language teaching (any language, not just English!), teacher training, managing schools or developing language learning materials can benefit from using the framework.
How to Make a Field invisible in Odoo 17Celine George
It is possible to hide or invisible some fields in odoo. Commonly using “invisible” attribute in the field definition to invisible the fields. This slide will show how to make a field invisible in odoo 17.
Instructions for Submissions thorugh G- Classroom.pptxJheel Barad
This presentation provides a briefing on how to upload submissions and documents in Google Classroom. It was prepared as part of an orientation for new Sainik School in-service teacher trainees. As a training officer, my goal is to ensure that you are comfortable and proficient with this essential tool for managing assignments and fostering student engagement.
Model Attribute Check Company Auto PropertyCeline George
In Odoo, the multi-company feature allows you to manage multiple companies within a single Odoo database instance. Each company can have its own configurations while still sharing common resources such as products, customers, and suppliers.
2. CONTENT
INTRODUCTION
HISTORY OF CRYOPRESERVATION
OBJECTIVE OF CRYOPRESERVATION
MILT COMPOSITION AND SPERM QUALITY
OF TELEOSTS
PRINCIPLE OF CRYOPRESERVATION
CRYOPROTECTANTS
EXTENDERS
DILUENTS
3. CRYOPRESERVATION PROTOCOL
FACTORS AFFECTING THE POST THAW
FERTILITY OF CRYOPRESERVED
SPERMATOZOA
ADVANTAGES OF CRYOPRESERVATION
OF GAMETES
4. Cryopreservation is the ex-situ conservation of
fish spermatozoa, eggs and embryos in ultra
low temperature such as -196°C
cryopreserve the spermatozoa and ova from
various species and to use the same for seed
production
Cryopreservation of female gametes of fish
could not be done successfully
In india, the NBFGR is the primary
organization carrying out fish sperm
cryopreservation
5. The feasibility of preservation of living cells in super
cold temperature
Cryopreservation was first demonstrated by polge et
al in 1949
This had stimulated extensive cryobiological
research
J .h .s. blaxter first successfully cryopreserved the
spermatozoa of herring in 1953
In india 27 species for sperm cryopreservation are
prioritized
The NBFGR has initiated research programme on
cryopreservation of blastomeres of catfishes.
6. Germplasm conservation
Stock improvement for better growth
Sex synchronization for research
Desirable quality, quantity of gametes
Available of disease resistance and pathogen free
seed
sperm cryopreservation protocol can be an asset
where such milt related problem asynchronization
exsist
The sperm cryopreservation has also been tested
for production of rohu seed
7. Milt is the seminal fluid/seminal plasma
containing mature spermatozoa
Seminal plasma is the source of nutrition for
the spermatozoa.
Seminal plasma also inhibits sperm motility
Motility of spermatozoa of salmonids last only
up to 2 minutes
Once the fertility is lost, they loss there
fertilizing ability too
Seminal plasma which is secreted by the
secretory cells lining
8. Seminal plasma contains both organic and
inorganic components
Monosaccharides and lipids are considered
essential for the nourishment of mature
spermatozoa
Presence of K+ in the seminal plasma has been
shown to inhibit flagellar movement/motility
of the sperm
Presence of citric acid in seminal plasma is
crucial to keep the sperm cells inactive in the
sperm duct
In the seminal plasma, calcium and magnesium
are found bound to citric acid
9. A few parameters have been developed to assess the
quality of sperms for cryopreservation
Spermatocrit value
Sperm density
Milt volume
Motility of spermatozoa
Age of spermatozoa
For the milt of carp, spermatocrit value of 70 and above
is considered good
In case of indian major carps sperm density may vary
from 2×10⁷ to 3.5×10⁷ per cu.mm
If percentage of spermatozoa exhibiting movement is
about 70% or more it is considered good
For cryopreservation, milt from the middle of spawning
season has been shown to be good
10. There are three essential steps in cryopreservation
Freezing
Storing
Thawing
The injuries to the cells may be due to
Formation of ice crystals during freezing and thawing
Due to osmotic changes
The rate of freezing and rate of thawing are critical in
preserving the vitality and viability of the spermatozoa
Moderate freezing is advisable
If the cooling rate is too low, due to dehydration the cells die
If the cooling rate is very high, the cells equilibrate
by intracellular freezing either by homogeneous heterogeneous
nucleation
The formation of large intracellular crystal is fatal for the cells
11.
12. Vitrification is the process that transforms
intracellular water to noncrystalline solids after
freezing
This occurs under two circumtances;
If the cooling rate is very high, it does not allow
sufficient time for the water the water molecule to
crystallize
The solution is so concentrated that the high
viscosity at low temperature does not allow water
molecule to get crystallize
The temperature at which vitrification begins is
called glass transformation temperature
Glass transformation temperature -13°C for water
13. Cryoprotectants are chemicals that minimise cryoinjuries
to the cell due to ice formation or suppress ice formation
The use of cryoprotectants increase the rate of
vitrification and optimise the post –thaw survival
Cryoprotectants are of two categories
Permeating cryoprotectants
Non permeating cryoprotectants
PERMEATING CRYOPROTECTANTS
Cryoprotectants that are permeable
to cell membrane
Dimethyl sulfoxide (DMSO)m
Glycerol,
Methanol
1,2 propanediol or propylene glycol
14. It should be least toxic to the cells
It should be permeable to the cells
It should be soluble in water during freezing
DMSO is the most commonly used cryoprotectants
Methanol which is considered most toxic
Glycerol is least toxic
Though glycerol is least toxic, it is less permeable to
the cell membrane
15. These chemicals are not permeable to cell membrane
The commonly used cryoprotectant are;
Sucrose
Glucose polymers such as ;
Dextran
Hydroxyethylstarch
Poly vinyl pyrrolidone(PVP)
Egg yolk serum
Skim milk
Anti freeze protein
They act by depressing the freezing point and raising the
glass transformation temperature of extracellular solution
cryoprotectants plays the role of dehydrant, freezing point
depressant and extra/intra cellular stabilising agent
16. It is a solution of balanced salts
It is also inhibits the activation of spermatozoa
This solution is meant to dilute milt as undiluted
milt is not suitable for freezing
It functions as medium for cryoprotectant
Usually, the diluents used for milt is the
combination of extenders and cryoprotectants
For Indian major carps, 3 categories of extenders
have been found suitable
18. Diluents is the combination of extenders and
cryoprotectants
Three categories of diluents are prepared by mixing
cryoprotectants
ǀ ǁ ǁǀ
Extender, A -90ml Extender, B -90ml Extender, C -65ml
DMSO -5ml DMSO -8ml DMSO -15ml
Glycerol -10ml Glycerol -10ml
Diluents can be mixed with milt at a proportion of 4:1 (
Diluents:Milt )
19. It can be described in 5 phases
Prefreezing
Freezing
Storage
Thawing
Post thaw and insemination
PRE-FREEZING PHASE
Collection of milt from the milter
Equilibration of milt diluents mixture
The milt diluents mixture is kept at low
temperature for equilibration
20.
21. FREEZING PHASE;
The equilibrated milt –diluents mixture is
frozen at low temperature
There are three ways of freezing viz;
Pellet method
It is extensively used for salmonids sperms
Size of the pellet may vary from 50 to 200ml
Straw method
Straws are available in different volumes (0.25
to 4ml)
This method is more popular during recent
years
22. Ampoules/vials/ visitubes method
STORAGE PHASE;
Frozen milt (pellet/ vial/ straw) is stored at
-196 °C
Stored in liquid nitrogen, kept in cryocans
THAWING PHASE;
For sperms frozen by pellet method, a thawing
solution is required
One pellet may be placed in 1-2 ml of thawing
solution
The straw are usually thawed in water bath at
30-40°C.
23.
24. Fast thawing is preferred as slow thawing may
cause crystallization
In case of ampoules/ vials 65-70 seconds are
required for slush formation
POST THAW PHASE AND INSEMINATION;
Sperm that have survived the cryopreservation are
ready for artificial insemination
Artificial insemination is to be done immediately
after thawing
Artificial insemination is to be done immediately
after thawing as there is reduction in the motility
period of spermatozoa after thawing
26. Phase Factors affecting
post thaw fertility
Prefreezing phase; Quantity of donor males(age, size, health)
Age of sperm and quality of milt
Type of concentration of diluents
Equlibration time and temp.
Interval between sperm collecting and freezing
Freezing phase; Freezing rate
Cryoprotectant/ extender used
Diluents preparation and dilution rate
Type of freezing container(pellet/ straw/ ampoule)
27. Storage phase; Temperature
Back ground radiation
Thawing phase; Thawing solution in case of pellets
Thawing rate
Motility induction and dilution
Removal of cryoprotectants
Post thawing &
insemination;
Sperm density for insemination
Quality of donor female for eggs
Incubation
28. It helps in the selective breeding
It can help in preventing inbreeding depression
It will be helpful in preserving genetic diversity
This procedure shall be useful in case of fishes
where male and female mature at different
time
It will be helpful in conserving endangered
species
It can reduce the cost of male brood stock
maintenance
29. Handbook of fisheries and aquaculture
Dr.S.Ayyappan
Breeding of fin fish and shell fish
P.C.Thomas
Applied fish genetics
B.K.Padhi
R.K.Mandal
Google.nbfgr( encyclopedia )
Google.wikipedia