M.K. Thong^1*, Z. Rudzki² ,J. HaIl², S. F. Yap³, KL. Tan³ and J.A.M.A. Tan³

¹Department of Paediatrics and ²Department of Allied Health Sciences, Faculty of Medicine,
University of Malaya, 50603 Kuala Lumpur, Malaysia . ³The Molecular Pathology Unit,
Institute of Medical and Veterinary Science, Frome Road, SA 5000, Australia.

(Received 7 December 1998 / Accepted 12 February 1999)

Abstract.
Advances in molecular diagnostic techniques have resulted in the characterisation of over 200 mutations causing ß thalassaemia. In addition, each population appears to have its own unique set and frequency of ß globin gene mutations. The strategy for prenatal diagnosis therefore involves screening at risk couples for the most common mutations known to be present in the ethnic group or population. As a result of this selective approach, uncommon or novel mutations are not detected. We undertook to characterise B globin gene mutations which remain undetected following polymerase chain reaction (PCR) and amplification refractory mutation system (ARMS) analyses. In this study, we successfully identified 13 of the 16 'unknown' alleles using chemical cleavage of mismatch (CCM) method and direct sequencing. As a result of this coupled strategy (ARMS and CCM), 98.5% of the ß globin gene mutations in the Thalassaemia Registry, University Hospital Kuala Lumpur were characterised.

Keywords: ß thalassaemia, chemical cleavage of mismatch, uncommon mutations, mutation detection

T.H. Tang¹, M. Musa¹, Z.F. Zainuddin¹, K.E. Choo²,L. Mohd Noh¹, K A. Myat¹,
1. Yaacob¹, H. Ismail², M. Mustaffa³ and Maheran Musa²

¹School of Medical Sciences, Universiti Sains Malaysia Cawangan Kelantan, Kota Bharu, Kelantan, Malaysia;
²Paediatric Unit, Kota Bharu Hospital, Kota Bharu, Kelantan, Malaysia; 3Chest Clinic,
Kota Bharu Hospital, Kota Bharu, Kelantan, Malaysia

(Received 8 December 1998 / Accepted 26 February 1999)

Abstract.
Tuberculosis (TB) in children is severe especially for those under 5 years old, with a tendency to extrapulmonary spread to other organs with high morbidity and mortality. Thus, in contrast to most adult cases, TB in children often requires urgent treatment but diagnosis is often difficult to establish by clinical and conventional laboratory methods. Rapid tests based on molecular techniques for diagnosis of TB have been widely reported but relatively few involve paediatric cases. In this study, a polymerase chain reaction (PCR) based test using primers derived from the insertion sequence IS 6110 and a new Mycobacterium tuberculosis complex specific target designated as B9 was developed and used for detecting the organism in specimens taken from four paediatric patients. This method was found to be a useful addition to the current battery of tests available to the paediatrician particularly in cases where diagnoses are difficult to establish.

Keywords: B9, IS6110, Mycobacterium tuberculosis, paediatric, PCR

Z. Zainal*, G. A. Tucker and G. W. Lycett

Department ofPhysiology and Environmental Science, Faculty of Agricultural and Food Sciences, University of Nottingham, Sutton Bonington Campus, Loughborough, Leicestershire. LE12 5RD, UK

(Received 15 February 1999 / Accepted 28 May 1999)

Abstract.
A full length cDNA clone designated as pNY601 encoding I aminocyclopropane I carboxylate (ACC) oxidase was isolated from a ripening mango (Mangifera indica L.) cDNA library using pTOM13 as a probe. Plasmid pNY601 is 1213 bp and contains a single open reading frame encoding a 324 amino acid polypeptide with a calculated molecular weight of 36.7 kDa and pI of 5.4. The deduced amino acid sequence shows considerable sequence (80-90%) similarity to other ACC oxidases; from various plant species. Northern blot analysis of RNA isolated from different stages of ripening shows that expression began while the fruits were still at the mature green stage. The appearance of these transcripts before ripening process take place may be due to chilling injury.

Keywords: Gene expression, Mangifiera indica, ACC oxidase, cDNA, fruit ripening

M.A. Abdullah¹, M. Marziah², M.N. Zakaria¹, A.M. Ali¹, N.H. Lajis³ and A.B. Ariff^1*

¹Department of Biotechnology, ²Department of Biochemistry and Microbiology, ³Department of Chemistry,
Universiti Putra Malaysia, 43400 UPM Serdang, Selangor, Malaysia

(Received 15 October 1998 / Accepted 10 May 1999)

Abstract.
Synergistic effects of medium strategies and sucrose levels with nitrogenous compounds, phosphate and myo inositol on Morinda elliptica cell suspension cultures were studied. Three types of medium strategies were tested maintenance (M), growth (G) and production (P) media. In M medium, incorporation of NO3 between 30 mM and 67.7 mM, was vital for optimum growth and anthraquinone (AQ) production. NO3 level upto 95.9 mM did not affect cell growth though AQ may be slightly reduced. At NO3 level of 77.1 mM, 8% (w/v) sucrose in G medium produced optimum cell growth and AQ content but cell growth was reduced whilst AQ content unaffected in P medium. At 10% (w/v) sucrose in both G and P media, both cell growth and AQ content were reduced. Phosphate was more a growth limiting nutrient than an AQ limiting nutrient. Absence of phosphate retarded cell growth in M medium but no significant difference on cell growth and AQ yield was observed with 5 fold increase in phosphate in M medium; at 8% (w/v) and 10% (w/v) sucrose in G medium; and at 8% (w/v) sucrose in P medium. At 10%(w/v) sucrose in P medium, the AQ yield was reduced by almost 50% (w/v). Nitrogen and phosphate toxicity was not only a function of both individual and collective level of ammonium, nitrate and phosphate, but also the level of sucrose and medium strategies. myo inositol was not an absolute requirement in M. elliptica cell suspension culture.

Keywords: Anthraquiriones, Morinda elliptica, nitrogen, phosphate, myo inosito