A technique for eliminating allele specific amplification failure during DNA amplification of heterozygous cells for preimplantation diagnosis

Advances in techniques of molecular biology have made possible the amplification of specific genes from single cells. This has a major clinical application in preimplantation diagnosis of monogenic disorders. However, the incidence of allele specific amplification failure (allele drop out) in heterozygous single cells can lead to misdiagnosis and the transfer of affected embryos. Few studies have been done to investigate the actual cause of allele drop out, although some investigators have succeeded in reducing but not eliminating it. Here we report the efficiency of amplifying both alleles in heterozygous cells lysed according to two different protocols. A total of 177 heterozygous cells from carriers of cystic fibrosis (CF) and haemoglobin C (HbC) were lysed using two different lysis buffers. Interestingly none of the cells that were lysed with sodium dodecyl sulphate/proteinase K showed any example of allele specific amplification failure whereas in those lysed by KOH/dithiothreitol it was present in 17.6 and 4.7% of the CF and HbC cells respectively. Our results suggest that the phenomenon of allele specific amplification failure is at least in part dependent on the lysis buffer used.      

Nomenclature for factors of the HLA system, update November/December 1996

The following sequences have been submitted to the Nomenclature Committee since the October 1996 nomenclature update and, following agreed policy, have been assigned official allele designations. Full details of all sequences will be published in the forthcoming report "Nomenclature for factors of the HLA system, 1996".
Below are listed the newly assigned sequences, confirmations of previously reported sequences and some sequences which are corrections of those originally reported. The accession number of each sequence is given and these can be used to retrieve the sequence files from either the EMBL, GenBank or DDBJ data libraries. Although accession numbers have been assigned by the data libraries and most sequences are already available, there is still the possibility that an author may not yet have allowed the sequence to be released, in such a case you will have to contact the submitting author directly.     

Some Actions of 9-Amino-1,2,3,4-Tetrahydroacridine (THA) on Cholinergic Transmission and Membrane Currents in Rat Sympathetic Ganglia

THA (Tacrine) is an anticholinesterase drug reported to alleviate cognitive deficit in Alzheimer's disease. We have used rat isolated superior cervical sympathetic ganglia as a model mammalian cholinergic neural system to study effects of THA on cholinergic synaptic transmission and postsynaptic membrane currents. At 0.1 - 3 microM, THA augmented the postsynaptic depolarizations and inward clamp currents produced by acetylcholine but not by the cholinesterase-resistant analogue, DMPP. Higher concentrations depressed these responses to both acetylcholine and DMPP, and reduced the acetylcholine-induced increase in membrane current noise. At 1 microM, THA did not affect the amplitude or time-course of fast (nicotinic) excitatory postsynaptic currents (epscs) evoked by single orthodromic volleys, but higher concentrations induced a biphasic epsc decay. In contrast, low concentrations of THA (1 - 3 microM) greatly augmented and prolonged the muscarinic slow epsc evoked by repetitive orthodromic volleys: this effect was blocked by 1 microM atropine. Concentrations above 0.1 mM produced a membrane depolarization and inhibited a variety of membrane ionic currents, including voltage-gated Ca current and subsequent Ca-activated K currents, and voltage-gated M- and A-type K currents. It is concluded that the principal effect of THA is to inhibit cholinesterase, and that the main consequence of this is to augment and prolong the muscarinic slow epsc. In contrast, the nicotinic fast epsc is not increased but instead may be reduced through a nicotinic channel-blocking action. Although THA could also block several other ion channels the concentrations required were too high to contribute significantly to its principal pharmacological actions on ganglionic transmission.     

When is enough enough? Secondary surgery for cleft lip and palate patients

Secondary cleft surgery differs from the primary surgery in two important respects: (1) the patient can participate in the choices of whether and how to proceed and (2) there is no universal consensus among professionals and lay persons alike that intervention is necessary. This article condenses a personal experience with secondary cleft surgery and attempts to answer "when is enough enough?"     

Tibial Plafond Fractures: Changing Principles of Treatment

Tibial plafond fractures from axial loading are high-energy injuries with significant associated soft-tissue damage. New classification methods include detailed anatomic subgroupings and highlight the soft-tissue injury. The traditional treatment of this intra-articular fracture with open reduction and internal fixation resulted in high rates of wound breakdown and infection. Treatment of these complications is lengthy and costly and not infrequently results in a poor outcome. Newer techniques using external fixation minimize disturbance of the soft-tissue envelope and have decreased these complications. Because the long-term outcome with all techniques is variable and often depends on factors beyond the surgeon's control, it is particularly important to avoid complications of initial treatment. Longer follow-up will determine whether patients treated with these techniques have a different rate of arthrosis.     

The biology of fracture healing: optimising outcome

Optimising the results of fracture treatment requires a holistic view of both patients and treatment. The nature of the patient determines the priority targets for outcome, which differ widely between the elderly and the young, and between the victims of high and low energy trauma. The efficacy of treatment depends on the overall process of care and rehabilitation as well as the strategy adopted to achieve bone healing. The rational basis for fracture treatment is the interaction between three elements: (i) the cell biology of bone regeneration; (ii) the revascularisation of devitalized bone and soft tissue adjacent to the fracture; and (iii) the mechanical environment of the fracture. The development of systems for early fracture stabilisation has been an advance. However, narrow thinking centred only on the restoration of mechanical integrity leads to poor strategy--the aim is to optimise the environment for bone healing. Future advances may come from the adjuvant use of molecular stimuli to bone regeneration.     

FIELD TRIAL OF Bacillus thuringiensis var israelensis PELLET FORMULATION IN THE CONTROL OF MOSQUITOES

In a simulated field trial Bacillus thuringiensis var israelensis (BTI) pellet formulation exhibited an enhanced efficacy with increasing dose. A dosage of 1.0 and 1.5 ppm was most effective under simulated field conditions. In field trials persistence of BTI pellet (1.0 ppm) was observed for 35 days in moderately polluted water collection as compared to 21 days in highly polluted water bodies.KEY WORDS: Bacillus thuringiensis, Malaria, Mosquito control