Gene ics
Selec ion
E olu ion
Tapio e al. Gene ics Selec ion E olu ion 2010, 42:28
h p://www.gsejou nal.o g/con en /42/1/28
Open Access
RESEARCH
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any medium, p o ided he o iginal wo k is p ope ly ci ed.
Resea ch
Es ima ion o ela edness among non-pedig eed
Yaku ian c yo-bank bulls using molecula da a:
implica ions o conse a ion and b eed
managemen
Ilma Tapio
1
, Miika Tapio
1
, Meng-Hua Li
1
, Ruslan Popo
2
, Zoya I ano a
2
and Juha Kan anen*
1
Abs ac
Backg ound: Yaku ian ca le, he las emaining na i e ca le b eed in Sibe ia, a e well adap ed o he ex eme sub-
a c ic condi ions. Nowadays only ca. 1200 pu eb ed animals a e le in Yaku ia. The semen o six Yaku ian bulls was
s o ed in a c yo-bank wi hou any pedig ee documen a ion because o he adi ional ee he ding s yle o he
popula ion.
Me hods: To cla i y he gene ic ela edness be ween hese bulls and o p o ide ecommenda ions o use hei semen
in u u e conse a ion and b eed managemen p og ams, we ha e analysed 30 au osomal mic osa elli es and
mi ochond ial DNA sequences in 60 indi iduals including he six o which semen has been s o ed. Fou ela edness
es ima o s we e calcula ed. In addi ion, we assessed he alue o he c yo-bank bulls o he p ese a ion o gene ic
a ia ion o he con empo a y Yaku ian ca le by calcula ing allelic and gene di e si y es ima es and mean molecula
coances ies.
Resul s: On he basis o mic osa elli e a iabili y, including he Yaku ian c yo-bank bulls inc eases he allelic a ia ion in
he con empo a y popula ion by 3% and in he male subpopula ion by 13%. In e ms o he mean molecula
coances ies, hey a e less ela ed o he con empo a y cow popula ion han he b eeding bulls and he e o e could be
used o educe inb eeding in he li ing popula ion. Al hough 30 loci a e insu icien o esol e de ini ely hei
ela edness ca ego ies, he da a sugges ou pai s o c yo-bank bulls as possible hal -sibs.
Conclusions: Ou esul s show ha e en ela i ely limi ed c yo-bank s o age o semen can ca y allelic a ia ion
h ough a bo leneck. We p opose a b eeding scheme based on he o a ion o b eeding emales and he di ision o
c yo-bank bulls in o h ee g oups. Thus, i molecula da a (e.g. au osomal mic osa elli e geno ypes) o he
con empo a y popula ion a e a ailable and based on ela i ely small-scale labo a o y analyses, i is possible o a oid
se ious mis akes in hei use o b eeding applica ions. The app oach sugges ed he e based on he use o Yaku ian
c yo-bank semen can be easily ex ended o c yo-bank ma e ials o o he animals in u u e b eeding p og ams.
Backg ound
Yaku ian ca le a e he las emaining na i e ca le b eed
o he Eas Asian 'Tu ano-Mongolian' ype o Bos au us
in Sibe ia. They a e dis ibu ed in he no h-eas e n
egion o he Sakha Republic (Yaku ia) o he Russian
Fede a ion [1-3]. These ca le possess a numbe o ai s,
such as solid unk, sho s ong legs and long hick win-
e coa , which make hem adap ed o he ex eme sub-
a c ic condi ions. Mo eo e , e icien he mo egula ion,
quick o ma ion o subcu aneous a y issue and low
me abolic a es a low empe a u es (e en down o -60°C)
allow hem o su i e in ha sh en i onmen s unde poo
eed condi ions (e.g. [3]). Ances o s o Yaku ian ca le can
be aced back o indigenous ca le in Sibe ia, which
mig a ed wi h he Yaku s ca. 1,000 yea s ago om he
sou he n Baikal egion o he no he n egions o he
Lena and Yana i e s. Yaku ian ca le we e pu eb ed un il
* Co espondence: juha.kan[email p o ec ed]
1 Bio echnology and Food Resea ch, MTT Ag i ood Resea ch Finland, Jokioinen,
FI-31600 Finland
Full lis o au ho in o ma ion is a ailable a he end o he a icle
Tapio e al. Gene ics Selec ion E olu ion 2010, 42:28
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1929 and, om hen on, we e subjec ed o ex ensi e
c ossb eeding wi h p oduc i e b eeds [2]. Consequen ly,
only ca. 1200 pu eb ed Yaku ian ca le indi iduals emain
in h ee illages in he dis ic o E eno-By an aisky, one
illage o Uluu-Syhyy and ou di e en a ms close o
Yaku sk Ci y [1]. Cu en ly he popula ion comp ises only
525 b eeding cows and 28 b eeding bulls. Yaku ian ca le
a e classi ied as an endange ed b eed by he Food and
Ag icul u e O ganiza ion o he Uni ed Na ions (FAO)
[4]. Howe e , ecen s udies in a con inen al con ex ha e
sugges ed ha his b eed is highly in e es ing o he con-
se a ion o ca le gene ic di e si y [3,5]. The e is a need
o conse e he b eed o u u e ca le b eeding ac ions as
well as o scien i ic and cul u al pu poses.
Main aining gene ic a iabili y and a oiding inb eeding
a e o g ea impo ance in he managemen o small ani-
mal popula ions. Inb eeding has a nega i e e ec on i -
ness, p oduc i i y and se e al o he pheno ypic ai s [6].
Meanwhile, a educ ion in gene and allele di e si y migh
educe a popula ion's esponse o en i onmen al changes
o a i icial selec ion in he u u e [7,8]. Thus, ex si u
banking o emb yos, oocy es and semen plays a unda-
men al ole in he conse a ion and managemen o small
a m animal b eeds [9]. S o age o gene ic ma e ial ep e-
sen s a ese oi o a b eed's gene ic di e si y and could
be used o e-es ablish a b eed, i needed. The only
gene ic ma e ial s o ed ex si u o Yaku ian ca le is he
semen om six bulls collec ed be ween 1980 and 1986.
Howe e , because o he adi ional ee he ding s yle o
hese ca le in summe pas u es, whe e se e al bulls ma e
andomly wi hin a he d, pedig ee eco ds o hese six
bulls a e no a ailable and, hus, he adi ional pedig ee-
based con ol o inb eeding is impossible in a meaning ul
way.
In he absence o pedig ee eco ds, molecula da a om
au osomal, ma e nally inhe i ed mi ochond ial DNA
(m DNA) o om pa e nally inhe i ed Y-ch omosomal
ma ke s can be used o es ima e ela edness be ween ani-
mals [10-12]. The widely applied s a is ical app oaches o
in e ela edness among indi iduals can be classi ied in o
wo ca ego ies: one in ol es he explici pedig ee econ-
s uc ion among all indi iduals in he sample; and he
o he is based on he bes pai wise ela ionship be ween
wo indi iduals a a ime based on ei he ela edness es i-
ma ion [13-15] o likelihood echniques [16,17]. The
weakness o he pai wise me hods is ha hey do no ake
in o accoun in o ma ion om he e e ence popula ion
and he di icul y in dis inguishing among ela ionships
wi h simila pa e ns o alleles (e.g. [18]). Howe e , pedi-
g ee econs uc ion me hods ha e been applied mainly o
he econs uc ion o ull-sib amilies [19].
Su i al o he las na i e ca le b eed in Sibe ia, Yaku-
ian ca le, is impo an o he local human communi y
as a sou ce o ood and income [1], bu also because i
p esen s ex eme adap i e po en ials o he ca le species
in gene al. Howe e , due o he small census size, Yaku-
ian ca le equi e a ca e ul managemen s a egy. Long-
e m c yo-conse a ion o emb yos and semen should be
conside ed se iously as hey ep esen a esou ce o
ongoing b eeding ac i i ies and a secu e way o p ese -
ing gene ic di e si y wi hin he b eed, should he li ing
popula ion encoun e p oblems. Al hough molecula
measu es o gene ic ela edness do no necessa ily ag ee
exac ly wi h he ue ela edness coe icien s calcula ed
om he pedig ee eco ds (bu see [20]), hey a e he bes
ela edness indica o s in he absence o eco ded pedi-
g ee in o ma ion (e.g. [11]). The e o e, he speci ic goals
o he cu en s udy we e o es ima e gene ic ela edness
among he six Yaku ian c yo-bank bulls using pai wise
and pedig ee econs uc ion me hods based on he analy-
sis o au osomal mic osa elli es and m DNA sequences.
We ha e also assessed how much gene ic a ia ion such a
limi ed ex si u bank could add o he con empo a y popu-
la ion o Yaku ian ca le. Ou aim was o sol e a p ac ical
conse a ion p oblem in a highly alued ca le b eed and
o see how help ul basic popula ion gene ics analyses a e
in sol ing such a b eed managemen ques ion. Ou
esul s also p o ide ecommenda ions o u u e conse -
a ion and use o he six c yo-bank semen.
Me hods
Sampling and da a ex ac ion
Genomic DNA was ex ac ed om he ozen semen
samples o six Yaku ian ca le c yo-bank bulls (named
Keskil, Moxsogol, Radzu, E el, Sa ial and Alii), whose
semen had been s o ed o mo e han 20 yea s, acco ding
o he me hod desc ibed by [21]. Fo he gene ic di e si y
compa ison, a e e ence popula ion consis ing o 54 an-
domly sampled Yaku ian ca le indi iduals om he S a e
a m in he illage o Kus u (17 indi iduals) and om
p i a e a ms in he illages o Ba agai-Aly a (17), Kus u
(4) and Uluu-Syhyy (16) in he Sakha Republic we e also
included in he analysis [3]. The e e ence popula ion
included samples o 37 cows and 17 bulls, e e ed he ea -
e o as ' he cow subpopula ion' and ' he bull subpopula-
ion', espec i ely. Geno ypes o he e e ence popula ion
using he same se o 30 au osomal mic osa elli es we e
ob ained om a p e ious s udy by Li e al. [3].
Molecula analysis
To de e mine he le els o m DNA a iabili y, DNA sam-
ples o he six Yaku ian c yo-bank bulls we e sequenced
o a 375-nucleo ide agmen o he m DNA con ol
egion using he p ime s published in [22]. The
sequenced agmen co e s bases 15,960 o 16,334 as
compa ed o he comple e ca le m DNA sequence
(NC006853). S anda d double-s anded sequencing was
pe o med wi h DYEnamic ET Te mina o Ki (Ame -
Tapio e al. Gene ics Selec ion E olu ion 2010, 42:28
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sham Biosciences) using he p ime s o polyme ase
chain eac ion (PCR) and 10 μL o pu i ied PCR-p oduc
on a MegaBACE™ 500 DNA Sequence (Ame sham Bio-
sciences). Complemen a y sequences we e combined
using he SEQUENCHER 4.6 so wa e (Gene Codes Co,
Ann A bo , MI, USA). In addi ion, sequences o 24 an-
dom indi iduals om he e e ence popula ion (accession
numbe s FJ014247-FJ014270) we e ob ained om a
ecen s udy [23]. The six Yaku ian c yo-bank bulls and
in e na ional e e ence animals we e geno yped o he
same se o 30 mic osa elli es (Table 1) as desc ibed in [3].
In o ma ion on p ime s and PCR condi ions can be
ound in he Ca le Di e si y Da abase h p://www.p oj-
ec s. oslin.ac.uk/cdi /ma ke s.h ml.
S a is ical analysis
To cha ac e ise he ma e nal lineages, mul iple align-
men s o m DNA sequences we e pe o med using Clus -
alX e sion 1.81 [24]. The size o he aligned m DNA
con ol egion agmen was 255 nucleo ides be ween
bases 16,021 and 16,275 compa ed o he comple e ca le
m DNA sequence (NC006853). The numbe o haplo-
ypes was es ima ed and pai wise gene ic dis ances
be ween haplo ypes we e calcula ed based on he numbe
o nucleo ide di e ences using MEGA e sion 3.1 [25].
Gene ic a iabili y o he au osomal mic osa elli e loci
in he whole Yaku ian ca le sample (60 indi iduals) was
quan i ied by he obse ed numbe o alleles (AO) and
polymo phism in o ma ion con en (PIC) pe locus using
he p og am Mic osa elli e TOOLKIT [26]. Locus-wise
es s o Ha dy-Weinbe g equilib ium (HWE) due o
he e ozygo e de iciency we e pe o med wi h 10,000
Mon e Ca lo andomisa ions [27] and he 'U' s a is ic es
[28] as implemen ed in he p og ams GENEPOP e sion
4.0 [29] and ML-Rela e [17], espec i ely. The p og am
GENEPOP was also used in he Fishe 's exac es s o
geno ypic linkage disequilib ium (LD) be ween all pai s
o mic osa elli es wi h a Ma ko chain me hod o 50,000
i e a ions and 100 ba ches.
Rela ionships among he six Yaku ian c yo-bank bulls
we e es ima ed wi h he pai wise ela edness es ima o s,
W [15] and QG [13], using he p og am SPAGeDi e sion
1.2 [30]. The calcula ion was based on au osomal mic o-
sa elli e geno ypes in all 60 indi iduals. Fu he mo e,
pai wise ela ionships be ween he bulls we e calcula ed
wi h he maximum-likelihood es ima o K using he p o-
g am ML-Rela e [17]. Pe o mances o W, QG, and K
we e e alua ed using a simula ion app oach as imple-
men ed in PEDAGOG [31]. Allele equencies o he 30
mic osa elli es ob ained om all 60 indi iduals we e used
as inpu da a. Dis ibu ion o pai wise ela edness (R)
es ima es o each o he ou simula ed ela ionship ca e-
go ies [un ela ed (UR), hal -sibs (HS), ull-sibs (FS), and
pa en -o sp ing (PO)] was based on he simula ed geno-
ypes om 1000 indi idual-pai s each. The sampling
a iance was calcula ed as he s anda d de ia ion o he
mean R es ima e o each simula ion ca ego y sepa a ely.
The bias among es ima o s was es ed by compa ing he
mean and he expec ed R alues (UR 0.0; HS 0.25; FS and
PO 0.5). Two- ailed - es s we e used o e alua e he sig-
ni icance o po en ial bias. C i ical signi icance alues
Table 1: In o ma ion on mic osa elli e ma ke s
Locus BTA AOPIC P
BM1824 1 4 0.39 0.547
BM2113 2 6 0.58 0.834
ETH10 5 5 0.39 0.543
ETH225 9 5 0.63 0.355
ETH3 19 4 0.60 0.908
HEL5 21 5 0.62 0.097
ILSTS005 10 3 0.32 0.813
INRA023 3 5 0.52 0.879
INRA035 16 3 0.18 0.002
INRA005 12 4 0.61 0.652
INRA063 18 3 0.33 0.407
BM1818 23 2 0.33 0.089
CSSM66 14 8 0.65 1.000
ETH152 5 4 0.51 0.969
HEL1 15 5 0.57 0.200
HEL13 11 4 0.44 0.143
HEL9 8 6 0.62 0.056
ILSTS006 7 6 0.67 0.115
INRA032 11 3 0.45 0.103
INRA037 11 5 0.52 0.265
TGLA227 18 8 0.68 0.421
TGLA126 20 4 0.68 0.216
TGLA122 21 6 0.67 0.745
HAUT24 22 4 0.50 0.747
HAUT27 26 8 0.69 0.389
CSRM60 10 7 0.62 0.206
MM12 9 4 0.51 0.181
ETH185 17 5 0.61 0.328
TGLA53 16 10 0.67 0.134
SPS115 15 4 0.41 0.264
Mean - 5 0.53 -
Fo each locus a e gi en he ch omosomal loca ion (BTA), and he
summa y s a is ics pe locus such as he numbe o obse ed alleles
(AO), polymo phism in o ma ion con en (PIC) and P- alues o he
de ia ion om Ha dy-Weinbe g equilib ium ac oss he o al 60
Yaku ian samples
Tapio e al. Gene ics Selec ion E olu ion 2010, 42:28
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we e adjus ed o mul iple es s wi h sequen ial Bon e -
oni co ec ion.
Pedig ee econs uc ion among all indi iduals in he
sample was pe o med using PARENTAGE e sion 1.0
[32]. Two chains wi h bu n-in o 200 i e a ions, hinning
o 400 and 2000 samples we e applied. A Di ichle p io
o he allele equencies was used and he p io o he
dis ibu ion o o sp ing be ween males and emales was
se o be gamma (1, 2). In luence o he six Yaku ian c yo-
bank bulls on he gene ic a iabili y o he e e ence pop-
ula ion and he bull subpopula ion we e in es iga ed by
calcula ing basic s a is ics such as gene and allelic di e si-
ies.
Molecula coances y is simila o he genealogical
coances y coe icien [33] bu is de ined as he p obabil-
i y ha wo alleles aken a andom, one om each indi-
idual, a e iden ical by s a e. To es i he six Yaku ian
c yo-bank bulls we e less ela ed o he cow subpopula-
ion (37 cows) han he bull subpopula ion (17 bulls), we
used he p og am MOL_COANC e sion 1.0 [34] o cal-
cula e he mean molecula coances y o he whole
Yaku ian ca le popula ion (60 indi iduals), o he all he
bulls (23 bulls comp ising he six c yo-bank bulls and he
17 e e ence bulls), o he 17 e e ence bulls, o six
Yaku ian c yo-bank bulls and o each o he 23 bulls sep-
a a ely. Mean molecula coances y [33] be ween each
bull and e e y cow was also calcula ed. The di e ence
be ween he bull subpopula ion (17 e e ence bulls) and
he g oup o six Yaku ian c yo-bank bulls was es ed
using a wo-sample pe mu a ion es by he Ho ho n and
Ho nik exac RankTes s e sion 0.8-12 package o he R
language.
Resul s
Mi ochond ial da a
Sc eening o he 255 n agmen o he m DNA con ol
egion iden i ied 11 haplo ypes de ined by 17 a iable
si es ha belong o he au ine m DNA sub-haplog oups
T2, T3 and T4 (Addi ional ile 1) [35,36]. Six haplo ypes
we e indi idual-speci ic, h ee haplo ypes we e sha ed by
wo samples, one haplo ype was sha ed by ou samples
and he mos common haplo ype was sha ed by 14 indi-
iduals. M DNA sequences o he six Yaku ian c yo-bank
bulls (accession numbe s FJ014464-FJ014469) we e cha -
ac e ized by six di e en haplo ypes, ou o which we e
no obse ed in he con empo a y samples (Addi ional
ile 1). The a e age numbe o pai wise nucleo ide di e -
ences among all 11 haplo ypes was 3.78, anging om 1
o 8 among pai s o compa ison. The numbe o pai wise
nucleo ide di e ences among he six haplo ypes obse ed
in he six Yaku ian c yo-bank bulls a ied om 2 o 7
wi h an a e age numbe o 4.53. We did no ind any
m DNA haplo ype sha ed by all six Yaku ian c yo-bank
bulls, which indica es ha hese bulls canno be ull-sibs
o ma e nal hal -sibs.
Mic osa elli es and ela edness
One hund ed and i y alleles we e de ec ed in he 60
Yaku ian ca le indi iduals ac oss he 30 mic osa elli es.
The numbe o obse ed alleles a ied om 2 o 10 pe
locus (Table 1). The a e age PIC ac oss he loci o he
comple e sample was 0.532, wi h he lowes PIC obse ed
a INRA035 (0.176) and he highes a HAUT27 (0.685).
No signi ican (P < 0.05) de ia ions om LE we e
obse ed in he pai wise mic osa elli e compa isons a e
sequen ial Bon e oni co ec ion was applied. Signi ican
(P < 0.05) he e ozygo e de iciency was de ec ed only a
INRA035 (Table 1), which is p obably due o he p esence
o non-ampli ying alleles (e.g. [37]). I is also possible ha
he locus INRA035 is nea a gene o wi hin a genomic
egion unde di ec ional selec ion and his would be
in e es ing o in es iga e u he .
We calcula ed pai wise ela edness es ima es be ween
he six Yaku ian c yo-bank bulls wi h and wi hou he
locus INRA035. These calcula ions o ela edness we e
u he adjus ed o accommoda e non-ampli ying alleles
by he op ion as implemen ed in he ML-Rela e p og am.
Nei he he exclusion o he locus no he inclusion o he
non-ampli ying alleles had a signi ican e ec on he
ela edness es ima es (no shown). The e o e, he esul s
p esen ed in he s udy a e based on he ull se o 30 mic-
osa elli es (Addi ional ile 2).
Mean W and QG es ima es and hei s anda d de ia-
ions calcula ed o ou simula ed ela edness dis ibu-
ions a e p esen ed in Addi ional ile 3. Pe o mances o
bo h pai wise ela edness es ima o s we e simila o each
o he wi h only mino di e ences in a iance es ima es.
QG had a sligh ly smalle (by 0.004) sampling a iance o
he dis ibu ion o UR indi iduals, while W pe o med
be e in he emaining ca ego ies (HS by 0.002, FS by
0.01 and PO by 0.015). In h ee ou o eigh cases, mean R
de ia ed signi ican ly om he expec ed alue (P < 0.013
a e sequen ial Bon e oni co ec ion). The bias o W
o UN pai s was downwa ds, while ha o W and QG in
he ca ego y o PO was upwa ds (Addi ional ile 3). The
pe o mance o K was e y simila o ha o W ( esul s
no shown) apa om nega i e alues being con e ed o
ze o ela edness.
Ten ou o 15 pai wise R-es ima es be ween he six
Yaku ian c yo-bank bulls app oached ze o o ell below i .
The emaining i e bull-pai s exhibi ed R- alues anging
om 0.124 o 0.276 o W and om 0.180 o 0.295 o QG
(Addi ional ile 2). All pai wise R alues we e plo ed on
he dis ibu ion o ou simula ed ela edness ca ego ies
(Figu e 1). When he W es ima o was used, one pai
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(Radzu:Sa ial, R = 0.276) ell ou side he 95% con idence
in e al o simula ed UR indi iduals ( he 95 h uppe
quan ile = 0.252) and was conside ed o be ela ed (Figu e
1a). Two o he pai s we e iden i ied as ela ed when he
QG es ima o was applied (Keskil:Moxsogol, R = 0.295;
Radzu:E el, R = 0.255; he 95 h uppe quan ile = 0.242)
(Figu e 1b). The ML-Rela e p og am uses simula ion o
de e mine which ela ionships a e consis en wi h geno-
ype da a and o compa e pu a i e ela ionships wi h
al e na i es. In o de o iden i y possible misclassi ied
indi iduals, a maximum-likelihood es ima o K es i-
ma ed by ML-Rela e was applied. Besides he h ee bull-
pai s men ioned abo e, he E el:Sa ial pai ( W = 0.205;
QG = 0.180) had he highes likelihood o being a hal -sib
Figu e 1 Pai wise ela edness o Yaku ian c yo-bank bulls. Values a e calcula ed using (A) W and (B) QG ela edness es ima o s plo ed on a dis-
ibu ion o ou simula ed ela ionship ca ego ies: un ela ed, hal -sibs, ull-sibs and pa en -o sp ing; he e ical line ep esen s he 95 h pe cen ile o
simula ed un ela ed indi iduals; he posi ion o pai wise alues in ega ds o he Y-axis was designed based on he es ima es om he K ela edness
es ima o and was calcula ed as 3 di ided by he cases when he log likelihood o R o he second closes ela ionship is smalle han he mos likely
ela ionship; abb e ia ions o Yaku ian c yo-bank bulls a e: K-Keskil, M-Moxsogol, R-Radzu, E-E el, S-Sa ial, A-Alii. Y-axis deno es he dis ibu ion o pos-
e io p obabili y densi y based on he simula ions o he ou ela ionship ca ego ies using he wo ela edness es ima o s W and QG, espec i ely.

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(Addi ional ile 2). The same ou pai s o Yaku ian c yo-
bank bulls we e also iden i ied as po en ial hal -sibs in he
pa en age analysis pe o med using he pedig ee econ-
s uc ion me hod among all indi iduals in he sample
(Addi ional ile 4).
Allelic di e si y and gene di e si y
Inclusion o he six Yaku ian c yo-bank bulls in he calcu-
la ion inc eases he wi hin-popula ion gene ic a iabili y
ela i e o ha in he con empo a y e e ence popula ion
(Table 2). Fo example, he six c yo-bank samples made i
possible o add wo new alleles a he locus INRA023 and
hei equency in he c yo-bank samples is 0.083. The e-
o e, compa ed o he h ee alleles de ec ed in he 54 con-
empo a y samples om he e e ence popula ion, a 67%
gain in allelic a ia ion was obse ed when including he
six c yo-bank samples. Wi h he six c yo-bank bulls, he
a e age allelic di e si y o he o al Yaku ian popula ion
inc eased by 3%, while he a e age allelic di e si y o he
bulls inc eased by 13%. F equencies o alleles speci ic o
he c yo-bank bulls anged om 0.083 o 0.250. Th ee
Yaku ian c yo-bank bulls, Keskil, Radzu and Alii, ca ied
alleles no de ec ed in he con empo a y Yaku ian popu-
la ion. Fu he mo e, all six Yaku ian c yo-bank bulls pos-
sessed mic osa elli e alleles ha we e no ound in he
con empo a y bull subpopula ion. The gene di e si y
would inc ease by 3.5% i he six c yo-bank bulls ep e-
sen ed he o al bull subpopula ion in he nex gene a ion
oge he wi h he con empo a y cows. The inc ease in
gene di e si y would be 1.2% by adding c yo-bank bulls o
he con empo a y bull subpopula ion in he calcula ion.
Molecula coances y
The mean molecula coances y was 0.416 o pai wise
compa isons among all 60 Yaku ian ca le indi iduals
(Table 3). The a e age molecula coances y calcula ed
be ween each Yaku ian bull and he cows anged om
0.344 o 0.465. Compa ed wi h he li ing con empo a y
bull subpopula ion, he g oup o six Yaku ian c yo-bank
bulls showed a signi ican ly lowe (0.395 s. 0.418; a pe -
mu a ion es be ween he wo mean alues, P = 0.035)
mean coances y wi h he li ing con empo a y cow sub-
popula ion (Table 3). This indica es ha he c yo-bank
bulls a e good candida es as si es in a b eeding p og am
aimed a a oiding inb eeding.
Discussion
Knowledge on pai wise ela edness is c ucial o d a ec-
ommenda ions o u he use o c yo-bank bull semen in
conse a ion and b eeding p og ams o domes ic ani-
mals. In his s udy, we ha e es ima ed pai wise ela ed-
ness among he six Yaku ian c yo-bank bulls wi h
di e en es ima o s based on au osomal and m DNA
gene ic a ia ion. Ou s udy has shown ha molecula
Table 2: Inc ease in allelic a ia ion when he six Yaku ian c yo-bank bulls a e included in he analysis
To al (N = 60) Males (N = 23)
NA% F equency Indi idual NA% F equency Indi idual
ETH10 0 0 - 1 25 0.167 Keskil, E el
HEL5 0 0 - 1 25 0.083 Sa ial
INRA023 2 67 0.083 Radzu, Alii 2 67 0.083 Radzu, Alii
INRA063 0 0 - 1 50 0.250 Keskil, Moxsogol, Sa ial
CSSM66 0 0 - 1 17 0.167 E el
ILSTS006 0 0 - 2 67 0.250 Keskil, Moxsogol
TGLA227 1 14 0.083 Keskil 1 17 0.083 Keskil
TGLA122 0 0 - 2 50 0.083 Moxsogol, E el
HAUT27 0 0 - 1 14 0.083 Keskil
ETH185 0 0 - 1 25 0.167 Keskil, Moxsogol
TGLA53 1 11 0.083 Keskil 1 17 0.083 Keskil
SPS115 0 0 - 1 50 0.083 Alii
A e age 3 13
The numbe (NA) and equency o added alleles in he six c yo-bank samples, he pe cen age gain in allelic a ia ion (%), and he name o he
Yaku ian c yo-bank bulls con ibu ing new alleles o he popula ion a e indica ed when all he samples (54 + 6 indi iduals) and he bull
samples (17 + 6 indi iduals) a e conside ed; he pe cen age gain in allelic a ia ion (%) was calcula ed by he numbe o added alleles in he
six c yo-bank samples di ided by he numbe o alleles in he 54 con empo a y animals om he e e ence popula ion
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da a p o ide a use ul ool o es ima e ela edness among
indi iduals when pedig ee da a a e una ailable. Mo e-
o e , he esul s clea ly demons a e he impo ance o ex
si u c yo-banking o gene ic ma e ial in he conse a ion
o a e domes ic animal b eeds.
Rela edness
Ou mic osa elli e analysis sugges s ha i e o he 15
pai wise ela edness compa isons o he Yaku ian c yo-
bank bulls exhibi ed coe icien s o ela edness (R) close
o he heo e ical expec a ions o hal -sibs (R = 25%) and
cousins (R = 12.5%). Howe e , he wo pai wise ela ed-
ness es ima o s iden i ied di e en Yaku ian bull-pai s as
clea ou lie s compa ed o he simula ed dis ibu ion o
andom indi iduals (Figu e 1). Rela edness es ima es o
simula ed un ela ed pai s ha e a e y wide dis ibu ion:
he 95 h pe cen iles ( QG = 0.242 and W = 0.252) a e e y
nea o abo e he heo e ical expec a ion o hal -sibs (R
= 0.25). This indica es ha he 30 mic osa elli e ma ke s
used he e a e insu icien o an unequi ocal sepa a ion
o ela ed and un ela ed indi iduals.
The numbe and gene ic a iabili y o ma ke s as well
as popula ion s uc u e migh a ec he obus ness o di -
e en me hods in he calcula ion o ela edness be ween
indi iduals. I has been also demons a ed ha he e is no
single bes -pe o ming es ima ion me hod o dis inguish
be ween all possible ypes o ela edness [38-40]. In his
s udy, W wo ked be e o he simula ed ca ego ies o
ela ed indi iduals ha a e impo an in sol ing ela ed-
ness ques ions among Yaku ian c yo-bank bulls. The
app oach by [15] is obus o a small sample size and in
he cases when he e e ence popula ion includes uniden-
i ied ela i es. These assump ions ma ch closely he si u-
a ion o he Yaku ian popula ion s udied and he e o e
could explain he be e pe o mance o he es ima o .
Thi y ma ke s seems o be su icien o iden i y PO's
o FS's, bu ails o sepa a e HS's o mo e dis an ela ed-
ness ca ego ies unequi ocally. Addi ional simula ions
ha e demons a ed ha a se o as many as 500 mic osa -
elli es would be needed o much mo e accu a e es i-
ma es o R wi h lowe s anda d de ia ions ( esul s no
p esen ed). Ou esul s ag ee wi h p e ious sugges ions
ha a la ge numbe o mic osa elli e loci a e needed o
unequi ocal cla i ica ion o pedig ees [33]. Al e na i ely,
using ad anced SNP-mic ochips wi h housands o SNP
could p o ide a solu ion (e.g. [41]).
Mi ochond ial da a
M DNA sequence analysis has shown ha he Yaku ian
c yo-bank bulls do no sha e any m DNA haplo ype.
Nucleo ide subs i u ions accumula e app oxima ely 5 o
10 imes as e in m DNA han in nuclea DNA [42] and
cases o m DNA mu a ion ixa ion wi hin one gene a ion
ha e been desc ibed in Hols ein ca le [43]. Howe e , he
smalles pai wise di e ences be ween haplo ypes
obse ed in Yaku ian c yo-bank bulls we e wo nucle-
o ides. As a esul o he e oplasmy, he sons o a dam can
ha e di e en m DNA haplo ypes. Howe e , no he e o-
plasmy was de ec ed in he p esen s udy. The m DNA
sequence analysis sugges s ha he e a e no ull-sibs o
ma e nal hal -sibs among he Yaku ian c yo-bank bulls.
Al hough ou Y ch omosome-speci ic mic osa elli es
(INRA124, INRA189, BM861 and BYM-1; see [44]) a e
monomo phic in he popula ion [23], he mean ela ed-
Table 3: A e age molecula coances ies
Indi idual Mean molecula coances y
To al popula ion 0.42
23 bulls 0.41
17 bulls 0.42
6 bulls 0.40
JA34 0.47
JA32 0.45
JA5 0.44
JA41 0.44
JA42 0.43
JA9 0.43
Alii 0.42
JA38 0.42
JA 17 0.42
JA44 0.42
JA33 0.42
Moxsogol 0.42
JA40 0.41
JA3 0.41
JA43 0.40
Sa ial 0.40
Radzu 0.40
JA11 0.40
JA39 0.40
JA31 0.39
E el 0.38
JA16 0.38
Keskil 0.34
Values a e calcula ed ac oss pai wise compa isons be ween he
indi iduals in he o al popula ion (60 indi iduals = 6 c yo-bank
bulls + 17 con empo a y bulls + 37 con empo a y cows) and he
con empo a y 37 cows, be ween he indi iduals in he o al bull
subpopula ion (6 c yo-bank bulls + 17 con empo a y bulls) and
he con empo a y 37 cows, be ween he bulls (17 indi iduals)
and he cows (37 indi iduals) om he Yaku ian e e ence
popula ion, be ween he six Yaku ian c yo-bank bulls and he
cows (37 indi iduals), and be ween each Yaku ian bull and he
cows (37 indi iduals) sepa a ely
Tapio e al. Gene ics Selec ion E olu ion 2010, 42:28
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ness based on he au osomal mic osa elli es shows ha
he e a e ou po en ial hal -sib pai s among he six Yaku-
ian c yo-bank bulls.
Allelic di e si y and gene di e si y
The six Yaku ian c yo-bank bulls appea o ep esen an
impo an sou ce o addi ional allelic a ia ion and gene
di e si y o he Yaku ian bull subpopula ion as well as o
he o al Yaku ian popula ion. A high le el o gene ic
di e si y would de e mine he i ness o indi iduals and
would a ec he po en ial esponse o a popula ion o
immedia e na u al o a i icial selec ion [45].
P ac ical ecommenda ions
In a small popula ion, misclassi ying ela ed indi iduals
as un ela ed ( ype II e o ) will esul in unde es ima ing
ela edness wi hin he popula ion and, hus, ep esen s a
isk o inc eased inb eeding a e in subsequen gene a-
ions. The e o e, we a e mo e conce ned abou minimiz-
ing he occu ence o ype II e o s a he han he
p esence o ype I e o s, whe e un ela ed indi iduals a e
iden i ied as ela ed. In he conse a ion p og am o he
Yaku ian ca le, we ecommend ha ou Yaku ian c yo-
bank bull-pai s, Keskil:Moxsogol, Radzu:Sa ial, E el:Sa -
ial and also Radzu:E el, a e ea ed as hal -sibs o indi id-
uals o he wise ha ing ela edness up o 25%.
In an endange ed popula ion, choosing op imal indi-
iduals o ma ing and designing an app op ia e ma ing
scheme can help o moni o he gene ic a ia ion and he
a e age ela edness among indi iduals. I has been shown
ha ma ing indi iduals wi h minimal a e age coances-
ies will maximize he popula ion's gene ic di e si y in
e ms o expec ed he e ozygosi y [46,47]. In ou s udy, 23
Yaku ian bulls a e candida e si es o he subsequen gen-
e a ion. Howe e , as compa ed wi h he con empo a y 17
bulls, he six Yaku ian c yo-bank bulls show signi ican ly
lowe a e age molecula coances ies wi h he cow popu-
la ion. Using he six c yo-bank semen in a i icial insemi-
na ion would help o con ol he a e o inb eeding in
ollowing gene a ions.
The choice o he ma ing sys em is complica ed
because o he ime scale o in e es . F om a sho - e m
pe spec i e, a simple b eeding scheme could be sug-
ges ed, whe eby a popula ion is subdi ided in o se e al
g oups and o a ion ma ing among hese g oups is pe -
o med [48]. In he o a ion ma ing scheme, b eeding
cows a e om he same g oup as he si e, while b eeding
bulls a e om a di e en g oup. Al hough his scheme
will no educe he deg ee o inb eeding in he long- un, a
mo e e en dis ibu ion o inb eeding among indi iduals
would be achie ed. Fu he mo e, i would gua an ee ha
each line p oduces p ogeny ha will be used o b eeding
in he nex gene a ion. On he basis o he pai wise ela -
edness among he six Yaku ian c yo-bank bulls, we sug-
ges o spli hem in o h ee sepa a e g oups in he
o a ion ma ing, wi h Alii alone in ano he g oup, Keskil
and Moxogol in one g oup, and Radzu, E el and Sa ial in
a hi d g oup.
Conclusions
Wi h he Yaku ian ca le as an example, ou esul s indi-
ca e ha e en a limi ed numbe o semen samples
selec ed o he long- e m c yo-banking can ep esen a
conside able po en ial o main ain wi hin-popula ion
gene ic a iabili y. The e o e, we ecommend en ichmen
o he c yo-bank by adding semen o un ela ed bulls wi h
new gene ic a iabili y om he cu en li ing popula-
ion. We ha e shown ha when pedig ee documen a ion
is una ailable, e en a limi ed numbe o molecula ma k-
e s can help o make e ec i e b eeding ma ing schemes,
hough a la ge se o ma ke s would be desi able. We
conclude ha he p esen s a egy wi h he help o molec-
ula da a can be applied o o he animal species o e en
plan s whe e he educ ion o inb eeding and he p ese -
a ion o gene ic a ia ion a e impo an conce ns.
Addi ional ma e ial
Compe ing in e es s
The au ho s decla e ha hey ha e no compe ing in e es s.
Au ho s' con ibu ions
IT designed he s udy, did he labo a o y wo k, pe o med he da a analysis
and d a ed he manusc ip . MT con ibu ed o he da a analysis and he d a
w i ing. MHL con ibu ed o he d a w i ing and e ised he manusc ip c i i-
cally. RP and ZY con ibu ed o he sample collec ion and he manusc ip w i -
ing and p o ided impo an expe ise on Yaku ian ca le. JK planned and
coo dina ed he whole s udy, and con ibu ed o he manusc ip w i ing. All
he au ho s ead and app o ed he inal manusc ip .
Acknowledgemen s
We a e indeb ed o A. Vi a o he echnical assis ance in mic osa elli e geno-
yping and m DNA sequencing, o V. Ahola o he help in bioin o ma ics and
o M. To o o his assis ance in coances y analyses. This wo k was pe o med as
a pa o he Russian in Flux- esea ch p og amme o he Academy o Finland
and was inancially suppo ed by he Academy o Finland. The o ice space
p o ided by ILRI in Nai obi, Kenya o I. Tapio a he inal s age o he s udy is
acknowledged.
Au ho De ails
1Bio echnology and Food Resea ch, MTT Ag i ood Resea ch Finland, Jokioinen,
FI-31600 Finland and 2Yaku ian Resea ch Ins i u e o Ag icul u e, 677002
Yaku sk, Sakha, Russia
Addi ional ile 1 Alignmen o he a iable si es in he 255 n ag-
men o he ca le m DNA con ol egion.
Addi ional ile 2 A e age ela edness es ima es o pai wise compa i-
sons among he six Yaku ian c yo-bank bulls ob ained using ela ed-
ness es ima o s W , QG and K.
Addi ional ile 3 Mean ela edness and hei s anda d de ia ions o
he wo ela edness es ima o s ( QG and W) o he ou simula ed
ela edness ca ego ies.
Addi ional ile 4 Resul s o he sha ed pa en age analysis.
Recei ed: 29 Ap il 2010 Accep ed: 13 July 2010
Published: 13 July 2010
This a icle is a ailable om: h p://www.gsejou nal.o g/con en /42/1/28© 2010 Tapio e al; licensee BioMed Cen al L d. This is an Open Access a icle dis ibu ed unde he e ms o he C ea i e Commons A ibu ion License (h p://c ea i ecommons.o g/licenses/by/2.0), which pe mi s un es ic ed use, dis ibu ion, and ep oduc ion in any medium, p o ided he o iginal wo k is p ope ly ci ed.Gene ics Selec ion E olu ion 2010, 42:28
Tapio e al. Gene ics Selec ion E olu ion 2010, 42:28
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