J Med Genet 1993; 30: 141-146

MEDICAL GENETICS AROUND THE WORLD

Molecular diagnosis of some common geneticdiseases in Russia and the former USSR:present and future

V S Baranov

AbstractThe current state of molecular diagnosisof some common genetic diseases,including cystic fibrosis, Duchenne mus-cular dystrophy, haemophilia A and B,phenylketonuria, and thalassaemia, inRussia and elsewhere in the formerUSSR is reviewed. Data on carrier detec-tion and prenatal diagnosis are presentedand some objective problems and ob-stacles hampering efficient moleculardiagnosis -in Russia are discussed. Thenecessity for molecular diagnosis ofsomeother inherited diseases (for example,von Willebrand's disease, Martin-Bellsyndrome, polycystic kidney disease,Huntington's disease, and myotonic dys-trophy) is stressed. The need for estab-lishing new diagnostic centres dealingwith the most common diseases, as wellas rare genetic diseases, is substantiated.Perspectives on the implementation ofnew molecular methods and new techni-cal approaches (preimplantation embryodiagnosis, fetal cells selected frommaternal blood) are briefly outlined.(J7 Med Genet 1993;30:141-6)

Institute of Obstetricsand Gynecology,Academy of MedicalSciences of Russia,Mendeleevskaya line3, St Petersburg199034, Russia.V S Baranov

The recent disintegration of the former USSRinto 15 independent states has caused tremen-dous disorganisation, not only in the economyas a whole but in all other fields of social life,including health services and medical geneticsin particular. The devastating consequences ofthis largely bloodless revolution for medicalgenetics stem primarily from pre-existingdrawbacks in the organisation of medical gen-etic services throughout the former giganticmultinational country. Monopolisation ofscience by two capital cities, Moscow andLeningrad (St Petersburg since September1991), in conjunction with a very unequal andgenerally rather low level of routine medicalgenetic counselling and laboratory diagnosis,can primarily be held responsible for this.An urgent programme of promotion and

improvement of public medical genetic ser-vices in the USSR was elaborated and offi-cially approved by the special session of ourMinistry of Health early in 1988. In spite ofdefinite progress in the screening of newborns

for some inherited diseases, organisation ofnumerous (about 115) local medical geneticscommittees, inter-regional medical geneticscentres (about 25), as well as the creation of anAll-Union Centre for Medical Genetics, incor-porating the former Institute of Medical Gen-etics and a new Institute of Clinical Geneticsin Moscow, modest funding and the absence ofhard currency still significantly hampers pro-gress in medical genetics throughout thecountry. Fortunately, since January 1989,scientists from the most advanced laboratoriesin the USSR (mainly from Moscow and Len-ingrad and a few from other capitals of theUnion Republics) have gained access to hardcurrency through a new State Scientific Pro-gramme (Human Genome), headed by aca-demician A A Baev, and launched by theAcademy of Sciences.

Participation in this programme during1989 and 1990 has provided at least somescientists with grants partly paid in hard cur-rency and thus has enabled them to acquirenecessary equipment and reagents indispens-able for molecular analysis. It is still too earlyto estimate the significance of our HumanGenome Programme as a whole, but its im-portant impact on medical genetic studies inthis country is quite obvious and has beenhighly appreciated. Over 50 different medicalgenetics projects were funded through theHuman Genome Programme in 1991.The principal goal of the present paper is to

give an up to date review of moleculardiagnosis for widespread genetic diseases inRussia and the former USSR, with specialemphasis on the major problems facing ourspecialists in medical genetics dealing withmolecular studies.As several different laboratories are usually

involved in molecular studies of the same com-mon genetic disease, each unit will be con-sidered separately, indicating the participationof each laboratory dealing with a particulardisease.

Cystic fibrosisMolecular studies of cystic fibrosis (CF) werelaunched in 1985 after the author's one monthvisit to Great Britain as a WHO student.Professor Bob Williamson provided recently

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discovered DNA probes which enabled South-ern blot analysis of RFLPs to be established,first at the Institute of Experimental Medicineand, since 1987, in a newly organised labora-tory for prenatal diagnosis of inherited diseasesat the Institute of Obstetrics and Gynecology,Academy of Medical Sciences, St Petersburg.The latter laboratory acquired the status of anAll-Union Centre for prenatal diagnosis of CFin 1989.

Patterns of allelic polymorphisms in CFhave been analysed by collaborative studies innative populations from six different regions ofthe European part of the former USSR. Signi-ficant differences were discovered in the Lith-uanian population (Met, D7S23) as well as thesouthern Slavonian populations of Ukraniaand Krasnodar (D7S23).'CF studies were intensified after identifica-

tion of the gene and the discovery of the majorAF508 mutation. According to our data thefrequency of AF508 in native populations fromthe European part of the country varies signi-ficantly (from 30% in Moldova to 69% inUkrania) being, on average, around 50% forCF patients in the north-west and central partsof Russia.2

Large scale testing of the AF508 mutationhas become possible after adopting the PCRtechnique for dried blood spots on filterpaper.3 Some other interesting innovations ofPCR techniques have been suggested by ESchwartz's group at the Institute of NuclearPhysics in St Petersburg.4

Pilot screening studies in St Petersburg (ESchwartz, Russian Centre for Molecular Dia-gnosis of Inherited Diseases) and in Ukrania(L Livshitz, Institute of Molecular Biologyand Genetics, Kiev) favour a relatively lowfrequency of AF508 among newborns (1:40).The other major mutations of the CFTR geneknown to be quite common in western popula-tions (G551D, R553X, R334W, W1282X,R551X, 1716+ 12T-*C) are detected only oc-casionally (1 to 3%) in our CF patients.5However, CFTR gene mutation 3732delA

(exon 19), recently found in southern France,was detected in almost 7% of Russian CFchromosomes in our collaborative studies withM Dean's laboratory (NIH, NCI, Frederick,USA).6 Moreover, 1677delTA, originally dis-covered by our group7 in Georgia (Megreliaregion) turned out to be a major mutation inthe populations of the Black Sea Basin. Threemore new CFTR gene mutations (E504Q exon10, W1282R exon 19, and S 1196X) have beenrecently shown by SSCP analysis followed bydirect sequencing.7According to these data the search for new

mutations of the CFTR gene in our CFpatients might be highly productive. At leastthree different mutations, AF508, 1677delTA,and 3732delA, might be used both for carrierdetection and prenatal diagnosis of CF in Rus-sia. These data are of special practical value asmost of the families requesting prenatal dia-gnosis of CF in this country do not have aliving index child and thus cannot be subjectedto RFLP analysis.

Around 200 prenatal diagnoses for CF havebeen carried out so far in the St PetersburgCentre. In 40 cases this relied exclusively onmolecular analysis, in 72 on molecular studiesplus microvillar enzyme testing, and in the rest(88 cases) only on the latter. A diagnosis of CFhas been made in 51 fetuses at risk and thesepregnancies were terminated.8

Molecular analysis of CF is carried out inclose collaboration with our centre in the Insti-tute of Experimental Medicine, Academy ofMedical Sciences, Laboratory of BiochemicalGenetics (phenotype-genotype correlations,RFLP analysis910), in the Russian Centre forMolecular Diagnosis of Inherited Diseases(pilot screening programme, intragenic poly-morphism studies), both in St Petersburg, inthe Centre of Medical Genetics, Moscow (mu-tation identification11), and in the Institute ofMolecular Biology and Genetics, UkraniaAcademy of Sciences, Kiev.12

Identification of new major CFTR genemutations, specific to our native populations,as well as RFLP analysis of new intragenicpolymorphisms, such as the recently discov-ered highly polymorphic minisatellite DNAsequences in introns 6, 8, and 17b, is helping inmore efficient application of molecular analysisin CF patients.As this centre still remains a single unit in

Russia, and even in the whole former USSR,for prenatal diagnosis of this very commongenetic disease, the foundation of new molecu-lar diagnostic centres dealing with high riskCF families is very urgent.

Duchenne muscular dystrophyMolecular diagnosis of Duchenne musculardystrophy (DMD) was started independentlyand almost simultaneously by two researchgroups, our laboratory in St Petersburg and 0Evgrafov's group in Moscow (now Laboratoryof Molecular Diagnosis at the Institute ofClinical Genetics).

Initial studies were carried out with intra-genic and flanking DNA probes, generouslydonated by Drs Kay Davies and T Monaco(UK) as well as by Drs L Kunkel and MKoenig (USA). These studies were later sup-plemented with multiplex polymerase chainreaction (MPCR) for exon deletion detectionin the dystrophin gene. Both carrier detectionand prenatal diagnosis have been done by thetwo groups. According to our collaborativedata,'3 DNA analysis has been provided for119 at risk families with at least one affectedchild, or with an affected close male relative ofa woman at risk of being a heterozygous car-rier. Most (about 80%) of the families wererepresented by a sporadic case. MPCR for 11different exons ascertained dystrophin genedeletions in 41% of our patients (49 out of thetotal 119). A relatively low deletion detectionrate with the standard set of exons tested byMPCR, a somewhat unusual pattern of dele-tion distribution along the DMD cDNA, and asignificant proportion of extensive deletionsextending through the major part of the gene,substantiated possible population differences

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Molecular diagnosis of some common genetic diseases in Russia and the former USSR: present and future

of DMD mutations in Russian patients com-pared with published collaborative data.'4More studies are already in progress both in

St Petersburg and in Moscow to ascertain or toreject possible population diversity in the pat-tern of common deletions in the dystrophingene and to study their population frequenciesand vulnerability of each exon during germcell proliferation and meiotic recombination.Of 94 families, 76 were found to be informa-

tive by RFLP analysis. Carrier status wasconfirmed in 12 and excluded in 21 femalerelatives in 21 DMD families. Out of 22 pre-natal diagnoses, eight affected fetuses werediagnosed prenatally by direct deletion testingor by RFLP analysis.Meanwhile, both carrier detection and pre-

natal diagnosis of DMD are still far from afinal solution in Russia. One of the possibleapproaches, not yet tried here so far, concernsRNA amplification supplemented with MPCRanalysis of cDNA. 5 Of special diagnostic valuefor at least some male fetuses at risk might alsobe the application of western blotting tomuscle biopsies or direct immunocytochemicalstudies of dystrophin in muscle fibres. Thisapproach might be of great benefit for theotherwise uninformative DMD familiesrequesting prenatal diagnosis during thesecond trimester of pregnancy.

Haemophilia AMolecular diagnosis of haemophilia A in Rus-sia was started in 1987 by our research groupin St Petersburg and by G Soloviev's group inthe All-Union Haematological Centre (Mos-cow). DNA probes were kindly provided byDr J-L Mandel (France) and Drs R Lawn andE Gitschier (USA). Over 200 at risk familieshave already been studied, most of them fromRussia (130) and the rest from haemophiliacentres in other states (Ukrania, Moldova,Belorussia, Uzbekistan etc).Highly polymorphic flanking DNA (probe

StI4/TaqI) and intragenic polymorphic sitesHindIII (intron 19), BclI (intron 18), andXbaI (intron 22) were used both for popula-tion studies and diagnostic purposes. Someintragenic polymorphic sites were used forpolymerase chain reaction detection.'6

Frequencies of allelic polymorphismsstudied for both intra- and intergenic probesin the Slavonian and Uzbekian populationswere found to be very similar and closelyresembled those in western European popula-tions. Two unusual alleles of the Stl4/TaqIsite have been discovered by our group in someSlavonian and Uzbekian samples.'7

Carrier status has been confirmed in 43 andrejected in 49 close female relatives of theproband in 1 13 families under our supervision.Haemophilia A has been diagnosed prenatallyin 18 out of 30 cases studied so far by ourgroup. Several prenatal diagnoses by PCRhave been carried out by the Moscow group incollaboration with their German colleagues.'8Mutation identification by SSCP analysis fol-lowed by direct sequencing of altered exons

and amplification-mismatch detection studiesof the factor VIII gene are now in progress.

Haemophilia BIntragenic DNA probes available for RFLPanalysis of the factor IX gene were kindlydonated to the author by Professor G Brown-lee (UK) in 1985. Southern blot RFLP analy-sis with these probes was later replaced byPCR for detection of intragenic polymorphicsites TaqI, XmnI (both in intron C), andHinfI/DdeI (intron A). Polymorphism identi-fication was done with original sets of oligo-primers suggested in collaborative studies withG Solovijv's group in Moscow.RFLP analysis of different native populations

has shown an unusual allele (most probably alarge insertion) in the HinfI/DdeI polymorphicsite in two Uzbekian DNA samples.

Carrier status has been proved in 10 andrejected in six female relatives of affected menin 18 families at risk. An unusually large dele-tion extending through both neighbouringexons G and H has been discovered in onepatient of Slavonian origin.

It should be mentioned that RFLP analysisis still informative in about 60 to 70% of at riskfamilies. Thus, direct identification of muta-tions by means of SSCP analysis or by theamplification-mismatch detection technique isadVisable. Both of these approaches are now inuse in our laboratory.

PhenylketonuriaLarge scale newborn screening programmesfor phenylketonuria (PKU), either by theGuthrie test or by an automated fluorescentassay, was officially recommended by the Min-istry of Health of the USSR in 1988. Accord-ing to already available data the frequency ofPKU in newborns varies between 1 in 5000 to1 in 8000 in different regions, with an averageof around 1 in 6000 (G Zuckerman, Instituteof Medical Genetics, Minsk, Belorussia).Molecular analysis of PKU in the USSR

was initiated with the cDNA probe of the PAHgene, generously provided by Dr X Woo(USA) to Dr V Kalinin (Laboratory of Mo-lecular Genetics, Institute of Medical Gen-etics, Moscow). These studies were substan-tially helped later by the PCR methodintroduced in Russia by Dr E Schwartz (Rus-sian Centre for Molecular Diagnosis of Gen-etic Diseases, St Petersburg). The fortunateavailability of thermostable DNA polymerasefrom Thermus thermophilis, manufactured bythe Institute of Nuclear Physics, Academy ofScience, USSR (St Petersburg) considerablyfacilitated a quick spread of the PCR methodthroughout the country since 1988.RFLP analysis of the PAH gene in Russian

Slavonians and in some other populations(Buryats) showed that the frequency of allelicpolymorphisms for the polymorphic sitesMspI, EcoRI, and HindIII is similar in thesepopulations and does not differ greatly fromother European populations.'9 New, efficientdetection of point mutations by the limited

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elongation method has recently been suggestedby this group in collaboration with Australianscientists.20

Allele specific hybridisation supplementedwith direct sequencing of exon 12 of thePAH gene showed the mutation in codon 408as well as the exon-intron splicing mutationto be predominant in Russian Slavonians(E Schwartz).More detailed identification of PKU muta-

tions in DNA samples from native USSRpopulations is now in progress in collaborativestudies of E Schwartz's Centre with X Woo'slaboratory at Baylor College (USA).2'

Eight of 12 families at risk for PKU werefound to be fully informative for the 408mutation in our collaborative studies with theCentre; 11 families at risk requested prenataldiagnosis and PKU has been confirmed inthree fetuses. These pregnancies were termi-nated.

P thalassaemiaThis genetic disease is confined to some south-ern republics of the former USSR, in particu-lar Azerbaijan, Uzbekistan, and Tadjikistan,where carrier frequencies are estimated to bearound 10% (G Solovjev). Molecular studiesof D thalassaemia were started in 1975 andwere initially confined to mRNA analysis andits application to deletion detection in the aglobin gene (L Lymborskaya). Detailed mo-lecular studies with more advanced techniqueswere undertaken by G Solovjev's group at theScientific Research Haematological Centrein Moscow and by E Schwartz's Centre inSt Petersburg. RFLP analysis in 32 e thalas-saemia patients with a severe form of thedisease showed 11 different haplotypes, one ofwhich was encountered in half of all affectedsubjects. This particular haplotype wasactually in strong linkage disequilibrium withmutations in codon 8. Five different mutationsof the globin gene were discovered in sevenpatients from Azerbaijan by PCR followed bydirect sequencing of the amplification pro-ducts. One of these mutations (deletion of a Gnucleotide between codons 82 and 83) wasfound for the first time.22 Oligoprimer setssuitable for allele specific amplification andthus for direct identification of i globin genemutations in the Azerbaijanian populationhave been suggested.23

Large scale screening of blood spots frompregnant women in Baku (Azerbaijan) hasrecently been undertaken and has resulted inthe identification of 66 at risk families (AKuliev, Institute of Medical Genetics, Mos-cow). Prenatal diagnosis of P thalassaemia hasbeen undertaken in 41 pregnant women and 14fetuses homozygous for D globin mutationsdiagnosed.

Wilson's disease (hepatolenticulardegeneration)While being quite infrequent (an average fre-quency in European populations of Russia isabout 1 in 20 000), WD deserves special atten-

tion as for many years it has been thoroughlystudied in the Laboratory of Biochemical Gen-etics, Institute for Experimental Medicine, inSt Petersburg (Professor S Nejfakh and hissuccessor Professor V Gaitzkhoki).

Identification and biochemical investiga-tions of copper binding protein caeruloplasmin(CP) in Wilson's disease patients were laterextended to CP mRNA and CP gene molecularanalysis.24 However, quite unexpectedly forthis group, these studies failed to disclose anymutations of the CP gene in patients and thusindicated that the CP gene is not involved byitself in Wilson's disease. The later assignmentof WD to chromosome 13 but not to chromo-some 3 (where the CP gene is located) agreedwith Neifakh's laboratory studies. Curiouslyenough, our in situ gene mapping with anoriginal fragment of rat CP-DNA as ahybridisation probe disclosed a positivehybridisation signal not only on chromosome 3(3q23-25, close to the transferrin gene) butalso on chromosome 13 (13q23-24), that is,somewhere very close to the still unknowngene responsible for Wilson's disease.25 Thusthe problem of the molecular nature of Wil-son's disease is still awaiting a solution.Meanwhile chromosome 13 is one of the

genome units selected by our Human GenomeProject for detailed molecular analysis. Identi-fication of the gene for Wilson's disease is oneof the urgent tasks for this project.

a, antitrypsin deficiencyDNA analysis of co, antitrypsin deficiency(AD) is confined to one laboratory (Laboratoryof Biochemical Genetics, St Petersburg, VGaitzkhoki) working in collaboration withCharles Coutelle's laboratory in Germany(Berlin). RFLP analysis was carried out in 659patients with chronic non-specific lung diseaseand a substantial preponderance of the abnor-mal Z allele was found (N Monakhov). Anunusual neutral mutation of the MaeIII site inexon 3 of the AT gene in 20 to 30% of normalsubjects with the MI allele, as well as itslinkage disequilibrium with the Z allele in ADpatients, was discovered. An allele specificamplification system for direct detection of thecommon mutation in codon 342 ofAD patientswith the Z haplotype was elaborated and tested(A Schwartzman).

Familial hypercholesterolaemia andother lipoprotein disordersMolecular analysis has been confined to 20families with familial hypercholesterolaemia(A Schwartzman). No major mutations or re-arrangements in the low density lipoproteinreceptor gene have been reported so far. RFLPanalysis of different polymorphic sites of thereceptor gene in normal and affected subjectsis in progress.

Correlation of particular genotypes of ApoCIII, Apo B 100, and Apo Al genes with bloodcholesterol lipoprotein levels in patients withcardiac ischaemic disease and in the generalpopulation is being studied by E Schwartz's

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Centre in St Petersburg26 and N Mertvetzov'sgroup in the Institute of Bio-organicChemistry in Novosibirsk. A simple approachbased on PCR mediated site directed mutage-nesis has been suggested for the identificationof the common mutation in codon 3500 of theapo BiO gene.27

ConclusionsAs might be inferred from these data, there arenow at least seven different laboratories inRussia dealing with the molecular analysis ofaround 10 different common genetic diseases.Many more genetic diseases have recently

become amenable to molecular studies bythese groups owing to the generous donationof new DNA probes, while even more geneticdisorders can be studied by the much moreefficient and convenient PCR techniques.Good candidates for molecular studies inRussia include the following diseases: fragile Xsyndrome, polycystic kidney disease, myotonicdystrophy, von Willebrand disease (St Peters-burg), Friedreich's ataxia, spinal muscularatrophy, Huntington's disease, some muco-polysaccharidoses, adrenogenital syndrome(O Evgrafov's laboratory at the Institute ofClinical Genetics in Moscow), in addition tosome forms of diabetes (E Schwartz's Centrein St Petersburg).Of these diseases fragile X mental retard-

ation is of special practical and scientific inter-est. Application of new DNA probes gener-ously provided by Drs Kay Davies and J-LMandel have opened the way for efficientcarrier detection and prenatal diagnosis ofthis syndrome. There are already over 20 dia-gnosed fra (X) families in St Petersburg andabout 70 in Moscow. A programme of activeclinical and laboratory search for new at riskfamilies and their subsequent molecular analy-sis is already in progress.The problem of identification of at risk

families is urgent, not only for the fragile Xsyndrome but also for most of the diseasesmentioned above and reflects the overall lowlevel of medical genetic services in Russia.This stems both from inadequate medicalgenetic counselling and unsatisfactory (verylimited and often inefficient) laboratory dia-gnostic services. Conspicuous advances in themolecular diagnosis of some common geneticdiseases on the one hand and the rather primi-tive routine medical genetic services on theother, constitute an obvious paradox in medi-cal genetics in Russia today. As a result, manyat risk families already amenable to molecularanalysis remain unknown and cannot benefitfrom molecular advances, while others who arealready diagnosed are not properly informedabout the possibility of prenatal diagnosis andcarrier detection or cannot obtain proper treat-ment for their disease. Another consequence ofthis situation is overdiagnosis of some diseases,for example, cystic fibrosis, which results in aflood of blood spotted filters sent to our Centreonly for the purpose of checking for the AF508mutation.Another major problem, closely linked with

the former one, is the very limited applicationof large scale screening programmes for PKU,galactosaemia, and hypothyroidism and a com-plete absence of newborn screening for CF(immunoreactive trypsin test).The absence of national and even local uni-

form regional registers of genetic diseasesaccessible for computer analysis is anotherserious obstacle to the active search and propercounselling of at risk families. Some promisingattempts to create corresponding automatedregisters of inherited diseases have beenundertaken in the Research Institute of Pae-diatrics and Child Surgery (Moscow), theInstitute of Medical Genetics (Moscow), andthe Institute for Physician Promotion andTraining (Moscow). An automated nationalregister incorporating information from manyrelevant local registers and accessible to medi-cal geneticists in molecular diagnostic labora-tories is an urgent need of medical geneticservices in this country.The clear advances in molecular studies of

genetic diseases in Russia, especially in recentyears, are largely the result of the close inter-national contact of the above mentioned medi-cal genetics groups with their colleagues inwestern Europe and the USA. The presentcritical political and economic situation in ourcountry, with its effects on medical genetics,cannot last forever. With the constant supportof our western colleagues and increasingopportunities for collaborative research pro-jects, medical geneticists in Russia will do theirbest to survive and to surmount this difficultperiod in our history with as few losses aspossible.

I wish to acknowledge our close and fruitfulscientific contacts with colleagues in the UK.Besides the names mentioned throughout thisreview, I am personally grateful to Professors JEdwards and E Southern (Oxford), ProfessorsM Bobrow and P Goodfellow (London), Pro-fessor M Ferguson-Smith (Cambridge), andProfessor Peter Harper, whose generous helpand constant interest in the work of my col-leagues and myself have had a major impact onadvances in molecular diagnosis of geneticdiseases in Russia and the former USSR as awhole. Wholehearted assistance of many otherprominent scientists in medical geneticsthroughout the world and especially in theUSA should also be mentioned. Financial sup-port of the Wellcome Trust to many of ourgeneticists, especially to 0 Evgrafov, V Mak-arov, and the author, is highly appreciated.

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2 Baranov VS, Ivaschenko TE, Gorbunova VN, et al. Fre-quency of the F508 deletion in cystic fibrosis patientsfrom the European part of the USSR. Hum Genet1991;87:61-4.

3 Schwartz EI, Khalchitsky SE, Eissensmith RC, Woo SLC.Polymerase chain-reaction amplification from dried bloodspots on Guthrie cards. Lancet 1990;316:639-40.

4 Scriabin VN, Khalchitsky SE, Kuzmin SE, Kaboev OK,Kalinin VM, Schwartz EI. A crude lysate of cells immo-bilized on solid support serve as matrix for enzymaticDNA. Nucleic Acids Res 1990;18:4289.

5 Baranov VS, Ivaschenko TE, Gorbunova VN, OsynovskayaNS, Gembovskaya SA. Five years experience in molecu-

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6 Ivaschenko TE, Baranov VS, Dean M. Two new mutationsand other CFTR gene molecular changes detected bySSCP analysis in CF-patients from Russia. Hum Genet (inpress).

7 Ivaschenko TE, White MB, Dean M, Baranov VS. Adeletion of two nucleotides in a Soviet family with cysticfibrosis causing early infant death. Genomics 1991;10:298-9.

8 Baranov VS, Gorbunova VN, Ivaschenko TE, et al. Fiveyears experience in prenatal diagnosis of cystic fibrosis inthe USSR. Prenat Diagn 1992;12:575-86.

9 Voronina 0, Gaitskhoki V, Potapova 0, Gembytskaya T.Genetic heterogeneity of clinical forms of CF. PediatrPulmonol Suppl 1990;5:202.

10 Voronina 0, Gaitskhoki V, Potapova 0, Schwartz E, Kir-ukhina L, Gembytskaya T. Correlation of F508 deletionwith CF clinical forms. Abstracts, 17th European CysticFibrosis Conference, Copenhagen, 1991:76.

11 Kalinin VN, Trubnikova IS, Barbova NI, Kapranov NI.DelF508 mutation in cystic fibrosis in Moscow region.Abstracts, 17th European Cystic Fibrosis Conference,Copenhagen, 1991:73.

12 Livshits LA, Kravchenko SA, Grishko VI, Gorovenko NG,Bujievskaya TI. Linkage disequilibrium between cysticfibrosis gene mutations and RFLP ofhuman chromosome7 within the Ukrainian population. Abstracts, 17th Euro-pean Cystic Fibrosis Conference, Copenhagen, 1991:74.

13 Baranov VS, Gorbunova VN, Malysheva OV, et al. Dystro-phin gene analysis and prenatal diagnosis of Duchennemuscular dystrophy in Russia. Prenat Diagn (in press).

14 Abbs S, Yau S, Clark S, Mathew CG, Bobrow M. Aconvenient multiplex PCR system for the detection ofdystrophin gene deletions: a comparative analysis withcDNA hybridisation shows mistyping by both methods. JfMed Genet 1991;28:304-11.

15 Roberts RG, Bentley DR, Barby TFM, Manners E,Bobrow M. Direct diagnosis of carriers of Duchenne andBecker muscular dystrophy by amplification of lympho-cyte RNA. Lancet 1990;336:1523-6.

16 Surin VL, Zhukova EL, Solovyev GY, Grineva NI. Simpleand convenient detection of a HindlII polymorphic site in

intron 19 of factor VIII using PCR. Nucleic Acids Res1990;18:3432.

17 Aseev MV, Malysheva OV, Baranov VS. Molecular analysisof some widespread X-linked diseases in different popula-tions of the USSR. Abstracts, 8th International Congress ofHuman Genetics, 1991:182.

18 Wehnert M, Surin VL, Zhukova EL, Solovyev GY, Herr-mann F. Prenatal diagnosis of hemophilia A by polymer-ase chain reaction (PCR) using Hind III polymorphism.Prenat Diagn 1990;10:529-32.

19 Kalinin VN, Laptev DA, Runkovskaya NG, Cotton RGH.Single base polymorphism in the coding region of thephenylalanine hydroxylase gene. Abstracts, EuropeanSociety of Human Genetics Annual Meeting, Leuven, Bel-gium 1991:99.

20 Kalinin VN, Ramus S, Cotton RGH. Detection of the pointmutations by the limited elongation reaction method(LERM). Abstracts, European Society of Human GeneticsAnnual Meeting, Leuven, Belgium 1991:175.

21 Woo SLC, Okano Y, Dasovich M, et al. Multiple foundingpopulations for phenylketonuria and prehistorical mi-gration in Europe. Proc Natl Acad Sci USA (in press).

22 Schwartz El, Goltzov AA, Kaboev OK, et al. A novelframe-shift mutation causing beta-thalassemia in Azer-baijan. Nucleic Acids Res 1989;17:3997.

23 Solovyev GY, Goltzov AA, Surin VL, et al. Molecularnature of mutations causing beta 0-thalassemia in Azer-baijan. Biomed Sci 1990;1:300-4.

24 Schwartzman AL, Gaitskhoki VS, L'vov VM, et al. Com-plex molecular structure of the gene coding for rat ceru-loplasmin. Gene 1980;11:1-10.

25 Baranov VS, Schwartzman AL, Gorbunova VN, et al.Chromosomal localization of ceruloplasmin and transfer-rin genes in laboratory rats, mice and in man by hybridi-zation with specific DNA probes. Chromosoma (Berl)1987;96:60-6.

26 Shevtsov SP, Kuchinski AP, Kovalev YR, et al. Geneticstudy of predisposition to myocardium infarction.Abstracts, 8th International Congress of Human Genetics,1991:203.

27 Schwartz El, Shevtzov SP, Kuchinskas AK, et al. Ap-proach to identification of a point mutation in apo-B100gene by means of PCR mediated site directed mutagene-sis. Nucleic Acids Res 1991;19:3752.

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