Bulletin USAMV-CN, 64/2007 (1-2).
MODERN METHODS OF DOGS’ KARYOTYPING
Pusta Dana*, Mariela Militaru**, Melinda Szalai***, I. Paşca*,
Rodica Sobolu*, Adel Ersek*
* University of Agricultural Sciences and Veterinary Medicine Cluj-Napoca, Faculty of Veterinary
Medicine
Str. Mănăştur nr. 3-5, 400372 Cluj-Napoca, Romania, e-mail: [email protected]
** University of Human Medicine “Iuliu HaŃieganu” Cluj-Napoca
*** Human Gynaecology I Clinic, Cluj-Napoca
Key words: dog, karyotype, MetaSystem Ikaros
Abstract. This study has the aim to realise the karyotype in dogs using new and original techniques of
staining, reading and realizing of the karyogram. To fulfill the proposed goal we used the classical visualization
techniques of the chromosomes in dog lymphocytes. We analyzed both metaphases and chromosomes in G
banding. The Karyotype was realized using a motorized Zeiss Axioscop 2 microscope having a capture camera
connected with a computer with a special software dedicated for the chromosomes karyotyping – MetaSystem
Ikaros. The software is normally used to karyotype the human chromosomes, but it was adapted by us to
karyotype the chromosomes in dog. Compared to the classical karyotyping methods, this software is very facile
and for this reason we recommend it to be used to realize the karyotype in animals.
MATERIAL AND METHOD
The method used to visualize the chromosomes was the classical one, using the cultures
of lymphocytes. The lymphocytes from the peripheral blood are grown in improved culture
media, treated by adding mitogens and, finally, the karyogram is made.
In the laboratory where we made the karyotyope in dog, to obtain the image we used a motorized Zeiss
Axioscop 2 microscope having a capture camera connected with a computer with a special software dedicated
for the chromosomes karyotyping – MetaSystem Ikaros
The blood used to for the leukocytes suspension was obtain from 10 adult dogs, five of them males and
five of them females. The results were obtained by reading and interpretation of 20 mitosis for each dog, by
reading both the mitosis itself and the G banded chromosomes.
RESULTS AND DISCUSSIONS
The MetaSystem Ikaros software normally used to karyotype the human chromosomes
and it was adapted by us to Karyotype the chromosomes in dogs (Fig. no. 1).
The MetaSystem Ikaros software has the possibility to receive and to process data captured by the Zeiss
capture camera attached to the Zeiss Axioscop 2 microscope, as follows:
Capture – receiving the image of a metaphase from the microscope
Add. capture – receiving an enlarged frame of an well spread metaphase image
Obj. Threshold – cleaning the image of the impurities on the examined slide
Mask Meta. – elimination of the lymphocytes in the image, for a better view of the
chromosomes
Delete – Deleting the impurities on the examined slide
Separate – separation of the chromosomes themselves (fig.11)
Overlaps – overlapping the chromosomes superposing
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Check Objects – checking the number of chromosomes
Annotate – annotation of the chromosomes in metaphase
Fig. 1 – Processing of the image captured from the examined slide
In Figure no. 2, we present the possibility to karyotype the chromosomes, after we
captured and “cleaned” the metaphase on the slide, using the same MetaSystem Ikaros
software.
Assign – optional arrangement, by the software, of the pair chromosomes
Rotate 180° / 90°, Rotate X° - rotation of the chromosomes for their correct
karyotyping
Shift – shifting the chromosomes to arrange them in pairs
Clean – “cleaning” the chromosomes
Reduce – reducing the image of the chromosomes
Magnify – magnifying the image of the chromosomes
Staining – staining, un-staining the G bans on the chromosomes
Annotate – annotation the chromosomes in metaphase
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Fig. 2 – Karyotyping the chromosomes in the captured metaphase
The final results of the dogs’ chromosomes karyotyping are presented in figures no. 3
and 4.
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Fig. 3 – The final result in male, after karyotyping
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Fig. 4 – The final result in female, after karyotyping
CONCLUSIONS
The method used to realize the karyogram was the cultivation of the lymphocytes from
the peripheral blood obtained from 10 adult dogs, 5 males and 5 females
The modern method of capturing, arranging and processing the chromosomes was the
MetaSystem Ikaros software, created for the human karyotype, but adapted by us for the
karyotype in dogs.
The MetaSystem Ikaros software is very easy to be used compared to the classical and
difficult karyotyping methods and we recommend it.
ACKNOWLEDGMENT
The authors want to thank to the doctors and especially to Mr. dr. Andrei Timen, PhD., from the TRIVET
S.R.L. Animal Hospital.
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