Mang N. et. al. Scientific Papers: Animal Sciences and Biotechnologies, 2011, 44 (1) [601158]

Mang N. et. al./ Scientific Papers: Animal Sciences and Biotechnologies, 2011, 44 (1)

435

The Establishment, Consolidation and Operation of Genetic
and Genome Banc of “Transilvanian Pinzgau” Breed in the
Hateg Land

Niculita Mang1, Ioan Vintil ă1, Alexandru Bogdan2, George Tob ă2, Florin Banateanu3,
Iudith Ipate2, Marcel Paraschivescu2

1West Univ. Vasile Goldi ș, B-dul Revolutiei nr. 94, Arad, Romania
2Rom. Acad. INCE-Center Res. for AgroFores. and Biodiv. Stud. Calea 13 Septembrie nr.13, Bucharest
3The Vet. Ag. for Healthand Food Safety, Bucharest

Abstract
Transilvanian Pinzgau breed is in danger of disappearing fr om the gene pool of cattle in Romania, massive crossings
currently practiced, even directed by the national breeding program. For this reason we have decided for now to
initiate a program of conservation, ex-situ by setting up a bank of genetic and genomic ,,Tara Hategului’’ led by the
Romanian Academy. The gene bank will preserve the biological mate rial containing ge rmoplasm population
composed and structured similar to that ex isting now in the breed, ecotype or line.

Keywords : genetic bank, Hateg, Pinzgau breed.

1. Introduction

Domestic animals species has a wide genetic
variety through which they can adapt to the
environment survive. Their genetic variation is
given, on the one hand, the multitude of breeds,
lines and ecotypes, who up the whole species and
on the other hand, the number of families and individuals that make up each breed component.
The more they are more numerous and more
diverse, with both genetic and species diversity or
breed will be higher and therefore they sustainable
time will be longer [1,2,3].
Genetic diversity of populations is eroded
Because some breeds of animals are more
important economically than others, many
traditional breeds, but are not economically
competitive are abandoned or absorbed into many
cross performances. Resu lt such a phenomenon is
not only a loss of cultural values national-
historical, but also a drastic reduction of genetic variability of the species, with negative consequences in its power to adapt to the environment [3,4,5]. Since the effect of amending
the genetic structure of character, because the
selection is greater, as the next generation involved in the production of fewer individuals, farmers are justified to use the reproduction of only 40-50% of the total herd of females (the
monotocics species ) and only 2% of all males,
whether artificial insemination is used. This results in a reduction, with each generation, the number of families in the breed and implicit effective population size (genetic size), making it impossible mating between unrelated individuals
only. This will result in: reduction of population
genetic variability is reflected in reduced
adaptability, population effect in increasing the
risk of genetic drift and the manifestation of
inbreeding depression drastic, especially for
characters contributing to the reproduction, the
vitality and survived at and increased disease
resistance [3,5]. All this obliges us to takes
conservation measures everyone, national genetic
resources. This means protecting ,,in situ’’ genetic
diversity in breeding programs or ,,ex situ’’ gene

Mang N. et. al./ Scientific Papers: Animal Science and Biotechnologies, 2011, 44 (1)

436banks by not only the most primitive breeds and
vulnerable or at risk of extinction, but also modern
and performing breeds [3,4,5,6].
Transilvanian Pinzgau’’ breed is in danger of
disappearing from the gene pool of cattle in
Romania, a massive crossings currently practiced,
even directed by the national breeding program.
For this reason we have decided for now to initiate
a program of conservation, ex-situ by setting up a
bank of genetic and genomic [7,8,9].

2. Materials and methods
Identifying animals that will produce biological
samples .
Before proceeding with sample collection, the
project team to create the gene bank to identify
families must pass pure breed component Transilvanian Pinzgau, Hunedoara region. It
should be based on pedigree, genetic markers, but also by information taken from OJRSA. After families have been identified and proven by
genomic analysis, donors of biological samples
are chosen, the record of this breed registration number, sex, age, owner, location, relevant phenotype, pedigree etc. The data obtained is entered in the data bank. It is proposed that the
Bank of genes in questi on are located in the
building Laboratory Biodiversity Studies Center of the Romanian Academy of General Common Berthelot Hunedoara county [1, 4, 9, 10].
Gene bank facilities
You must possess specific apparatus and
instruments for cell biology research laboratories,
embryology and embryo transfer, molecular
biology and a data bank. Institution staff is made
of genuine researchers. At least two of them, but
the manager must have higher education, highly
specialized in classical and population genetics,
embryology, cell culture, molecular biology and animal reproduction. Such knowledge is absolutely necessary, esp ecially in the use of
modern scholarship with specific genetic
laboratory, because even after the creation of the institution, especially after they have harvested samples to characterize the genetic and genomic, individual, family and breed and relationships other populations with whom they came in
contact. One of them must have sound knowledge
of working with your computer to can use national and international database. What kind of biological samples are needed?
To achieve the above objectives the following are
required:
-Cells : sperm in the ejaculate or the epididymis,
oocytes, embryos and soma tic cells, stem cells,
blood, skin, hair, ovaries and testes.
-Molecules : DNA and mitochondrial RNA, blood
All above mentioned types of cells, harvested
from Transylvanian Pinzgau addition that can be
preserved, always in liquid nitrogen, also have the
advantage that they can be used (except the
nucleic acids) in several ways to reconstruct
missing or when people get in danger of extinction
[3, 4, 5, 11]. -Sperm stored in liquid nitrogen after thawing can
be used to reconstruct the breed endangered or
extinct if they will be used to cross and ten back-
cross with females belonging to different genetic and other breeds that we call the supporting breed. Running back repeatedly for five generations with sperm collected from Transilvanian Pinzgau transformation genome will result in a very
helpful breed a race like we want to rebuild.
Genome progeny obtained after five generations of these crosses will contain a proportion of 94% genes from Pinzgau. [12, 13].
– Sperm after thawing, and immature oocytes
harvested in a certain way on the ovaries, after
thawing, followed by maturation can be user for fecundation, “in vitro'' in order to obtain viable embryos grown in the thermostat up to the blastocyst stage, when they can be transferred frozen to the uterine horns of surrogate mothers.
– Specialized somatic cells can be transformed
into embryos that are grown “in vitro'' in an enucleate oocytes. Oocyte cytoplasm has factors that are able to reprogram the genes in specialized cells and bring them to the embryonic state. An
electrical pulse, low intensity, has the gift to
induce entry into the soma tic cell division brought
in the state of the embryo and continue to embryogenetic and fetal development after its
transfer into the uterus of a surrogate mother. Such
a method helps us to multiply the population by
cloning. [5, 13].
– Pluripotent stem cells, or totipotent embrionic
stem cell have the advantage that can be use to
build artificial embryo coupling a group of some plural-potent stem cells, harvested Pinzgau with an embryos collected from another race but was
forced to become tetr aploid. Chimera will be
achieved through such technology will lose cells

Mang N. et. al./ Scientific Papers: Animal Science and Biotechnologies, 2011, 44 (1)

437during embryogenesis and tetraploid embryo will
become a pure-bred Pinzgau embryo [13]. Both specialized somatic cells and plural-potent stem cells longer the advantage that they can be mass used and cloning of unique genotypes found in
Pinzgau.
– Ovaries collected from Pinzgau breed and frozen in liquid nitrogen can be transplanted with the success, after thawing, another cow ovaries instead of another breed. They are functional and produce oocytes Pinzgau breed. It is understood
that such cows insemination with Pinzgau sperm,
defrost in the gene bank, will give rise only to pure-bred calves [3]. – Samples DNA and mitochondrial RNA used to assess genetic and genomic Pinzgau animals.
Analysis of (a micro-satellite and especially the
single nucleotide polymorphism-SNP haplo-types) will provide us with objective data on the size of genetic diversity of bree d, wealth alleles and
degree of heterozygous. With the SNP can determine the degree of erosion of genetic
diversity inbreeding and genetic drift. It can
establish the degree of breed division in ecotypes, or families in general and especially the level of immigration, inside it, the foreign gene race. Samples of DNA and mitochondrial RNA helps us
to establish the real extent of the genetic
population size (effective size – Ne), the genetic distance between Pinzgau breeds or the common origin of the breed with other breeds. Furthermore, molecular markers, help us to establish the objectivity of unique phenotypes, of excellent or
detrimental gene carriers and many other genetic
characteristics [5, 11].
How the collection of samples is made?
If the main purpose of ex-situ conservation of the breed is Pinzgau its full reconstruction in the
distant future, you will probably be in imminent danger of extinction or even disappeared, then we must take biological samples, on the whole, to gene-pool containing all of the breed. This means
that samples must be collected from all families
who make it up, because the number of families given the genetic structure of the breed or its actual effective size (Ne), by which time it can survive without too much erosion of its genetic diversity. If we take into account that family
members are very similar in genetic terms, that is
only enough to biological samples from one or two individuals within a family. Methodological guidelines for cryo-preservation
of genetic diversity of livestock breeds edited by FAO [12] recommended that a breed composed on 25 families, 20-30 different doses are sufficient semen and immature oocytes, collected from 25
bulls and all many cows. It is also recommended
that embryos and somatic cells for gene bank needed to be harvested from the same 25 males and 25 females which were collected and sperm and eventually. Samples of DNA and RNA-mt, and the blood serum, genomic characterization
user breed must be collected from more and
double the number of bulls and females. Are needed more evidence of such molecules to be
able to be used in rese arch to evaluate future
genomics, structure and genetic peculiarities.

3. Results and discussion

All samples collected fro m animals are identified
with a serial number and become the subject to specific processing of cells, tissue or molecules,
for freezing them in liquid nitrogen.

Types of collections of biological samples
Guidelines for ex-situ cryo-preservation of
domestic animal populations recommends that in
the gene bank should form several types of
collections of biological samples [3]. These are the followings: 1. Core collection It consists of all types of cells, tissues and molecules specified above and will be used rather in case of need of bred reconstruction,
after their disappearance or at the moment of
endangering by losing a high number of populations (bottleneck phenomenon). It should contain 150% of the samples needed for population reconstruction.
2. Working Collection Includes biological samples
identical to those of th e basic collection, and
samples taken from other animals with special genetic and phenotypic features, identified afterwards. From the working collection,
biological samples can be extracted for modern
genomic research to identify genetic markers of some singular or quantitative loci also called QTL. 3 Refreshing the Collection. Includes the same samples of cells, tissues and molecules that have been extracted from the wo rking collection, to be
examined, studied and pr ocessed, but need to be
refrozen after use.

Mang N. et. al./ Scientific Papers: Animal Science and Biotechnologies, 2011, 44 (1)

438Spatial organization of gene bank
Physical space will include at least the number of
rooms equal with the specific activities in the unit, and arranging them to follow a specific workflow. The activity of institution includes the following
components: a) collection of samples, b) specimen
processing, c) freezing, d) database, e) scientific research. Physical space will include at least the number of rooms equal with the specific activities in the unit, and arranging them to follow a specific workflow. Attention will be given maximum
spatial samples in liquid nitrogen, in order to
avoid accidents.
4. Conclusions
Germplasm or genetic pool of the Transilvanian
Pinzgau breed that will include representatives of biological samples collected from each family that compose the current population and stored in liquid nitrogen can be used in the following action:
1. The reconstruction of the whole bred
Transilvanian Pinzgau, where it will be necessary in the distant future, but to possess the same phenotypic and genetic characteristics as the current one. This is possible only if the preserved
biological samples will contain actual frequency
of genes and genotypes of the population, for all the characters that define it. 2. Minimize effects related mating and genetic drift, which certainly will occur in the future, the population intended to be pr eserved and ,,in situ’’.
3. The increase in population size genetic
(effective size) to increas e adaptive capacity and
to reduce genetic diseases population event that will be preserved in situ. 4. Biological samples containing the entire genetic
diversity of the breed will become a primary
source for genomics and proteomics research for discovery of new genetic markers for identification of genes and the interactions that underlie the manifestation of animal productivity,
the adaptability and disease resistance.

Acknowledgements

This work was co financed from the European Social
Fund through Sectorial Operational Programme Human
Resources Development 2007-2013, project number POSDRU/89/1.5/S/63258 ”Postdoctoral school for zoo
technical biodiversity and food biotechnology based on the eco-economy and the bio-economy required by eco-
sane-genesis”

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