Sheep gene mapping by somatic cell hybridization

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Sheep gene mapping by somatic cell hybridization

N Saïdi-Mehtar, M Imam-Ghali, S Heuertz, Mc Hors-Cayla

To cite this version:

Sheep

gene

mapping

by

somatic cell

_{hybridization}

N

Saïdi-Mehtar

M

Imam-Ghali

1

S

Heuertz

2

MC

_Hors-Cayla

₂

1

Université d’Oran

Es-Sénia,

Laboratoire de

_Biologie

M oléculaire et

_Génétique,

2 Unite

de Recherches de

_Biologie,

_Algeria;

z

INSERM

U12,

Unit6 de Recherches de

_{Genetique M6dicale, Paris,}

France

(Proceedings

of the 9th

_{European Colloquium}

on

_Cytogenetics

of Domestic

_Animals;

Toulouse-Auzeville,

10-13

July

1990)

sheep

/

ovine

_/

gene

mapping

/

synteny

/

cell _hybrid

INTRODUCTION

Twenty-five

hamster x

_{sheep hybrid}

cell lines were

_previously

obtained

(Saidi-Mehtar et

_al,

_1981b).

_Analysis

of these

_hybrids

has

_already

enabled the identification of 5

_syntenic

_groups and 10

_independent

markers

_{(Saidi-Mehtar}

et

_{al, 1981a, b, 1987;}

Millot et

_{al, 1981;}

Imam-Ghali

et

_al,

_1987).

In this _paper, we

_report

results obtained with 3 other _enzyme markers: aconitase 1

_(AC01),

inosine

_{triphosphatase}

_(ITPA)

and

_{glyceraldehyde-3-phosphate dehydrogenase}

_(GAPD);

and 6 DNA markers: ornithine

_{transcarbamylase}

_(OTC),

color vision red

_pigment

_(RCB),

_{raf oncogene}

(ARAF1)

and

_/3-gene

locus DR of human

_lymphocyte

_{antigen region}

_(HLA-DR!).

ENZYME

MARKERS

AC01,

ITPA and

GAPD

were studied

_by

cellulose acetate

_{electrophoresis using}

modified versions of the methods described

_by

Womack and Moll

_(1986).

The

cytoplasmic

(AC01)

and mitochondrial

_(AC02)

forms of aconitase were identified

by

successive

_freezing

and

_thawing

of cells followed

_by

_{electrophoresis}

of the

supernatants.

AC01

was obtained

first

and

_corresponded

to the most anodal band. A different

_{electrophoretic migration}

between

_sheep

and hamster was

only

observed

for

ACO1. Ovine

ITPA

_migrated

more

_anodally

than hamster ITPA.

Ovine GAPD

is a trimeric _enzyme:

_{positive hybrids}

for

_sheep

GAPD

showed 2 intermediate bands

_{(fig 1).}

_Comparison

between

_{AC01, ITPA,}

GAPD

and the other 21 markers

ITPA-adenosine

deaminase

_(ADA)

This

synteny

is conserved in man

(Hopkinson

et

al,

1976),

hamster

(Stalling

et

al,

1982),

mouse

(Siciliano

et

al,

1984),

cat

_(Berman

et

_al,

_1986),

cattle

_(Womack

and

Moll,

1986),

dog

and mink

_(Human

Gene

_{Mapping 9,}

_1987).

GAPD and lactate

dehydrogenase

B

_{(LDHB) peptidase}

B

(PEPB)-triose-phosphate

isomerase

_(TPI)

This _{group is syntenic only} in man

(Bootsma

and

Ruddle, 1978;

Ruddle and Meera

Khan,

1976),

cattle

_(Womack

and

_Moll,

₁₉₈₆₎

and mink

_(Human

Gene

Mapping

10,

1989).

This

_synteny

is not conserved in mouse, hamster and

rabbit,

where

an

_independent

_segregation

is observed between

GAPD-TPI-LDHB

and PEPB

(Human

Gene

_Mapping

_10,

_1989).

Like

_ADA,

ITPA is on U15. Like

LDHB-PEPB-TPI,

GAPD

is

assigned

to

chromosome

3.

AC01

_segregates

_{independently}

from the other 23

markers and

is

_assigned

to U2

_(Human

Gene

_{Mapping 10,}

_1989).

DNA MARKERS

DNA was extracted from the

parental

hamster cell

line,

ovine

lymphocytes

and

from 23 hamster x

_{sheep hybrid lines;}

3

_hybrid

lines were _grown

_{concomitantly}

blotting technique

using

the 6

_following

DNA

_{probes: OTC,}

isolated

_by

Horwich et

al

_(1984);

color vision red

_pigment,

isolated

_by

Nathans et al

_(1986);

one human

autosomal

_probe:

_HLA-DR,8,

isolated

_by

Wiman et al

_(1982);

two rat

X-linked

probes:

brain

_myelin

_{proteolipoprotein}

_(PLP),

isolated

_by

_Dautigny

et al

_(1985);

synapsin

1,

isolated

by

Kilimann and De

Gennaro

_(1985);

and one mouse X-linked

probe

ARAF1

_oncogene, isolated

_by

Huebner et al

_(1986).

Analysis

of

_hybrids

showed:

₁₎

the localization on the

_sheep

X chromosome

of sequences

homologous

to the

_following

_genes:

_OTC,

color vision red

_pigment,

ARAF1

_oncogene, brain

_myelin

PLP and

_synapsin

1

_(presence

of

molecular

_signals

in

_hybrids

_grown in HAT medium and absence of

_signals

in the same

hybrids

_grown in

_{6-thioguanine medium)}

_{(fig 2); 2)}

the absence of a

_positive

correlation between

the

_serological

_signal

obtained with anti-ovine

_{histocompatibility}

_complex

_(OLA)

sera

_(Nlillot

et

al,

1981)

and the molecular

_signal

obtained with the

_HLA-DR,8

In this

_study,

we showed two new

_syntenies

for the

sheep:

GAPD-LDHB-PEPB-TPI on chromosome 3 and ITPA-ADA on U15. The

_assignment

to the X chromosome of 5 genes, known to be X-linked in several other

_species,

confirms

the

_concept

of the conservation _{of the X chromosomal genome} in mammalian vertebrates.

ACKNOWLEDGMENTS

We are

_grateful

to AL

_Horwich,

J

Nathans,

A

_Dautigny,

LJ De

_Gennaro,

U

Rapp

and K Wiman _{for their generous}

_gifts

of

_probes.

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