Rapid prenatal diagnosis of aneuploidies and zygosity in multiple pregnancies by amniocentesis with single insertion of the needle and quantitative fluorescent PCR.

OBJECTIVE: To investigate amniotic fluid (AF) samples retrieved in multiple pregnancies by single insertion of the needle, for rapid assessment of chromosome copy number, zygosity, and cross-contamination between fetuses, using Quantitative Fluorescent Polymerase Chain Reaction (QF-PCR) amplification of highly polymorphic microsatellite markers. METHODS: Fifty-two multiple pregnancies were selected (47 twins, 5 triplets) and 108 samples of amniotic fluid were sampled between 12 to 20 weeks of gestation (mean 15.5) using the single-needle technique. Aneuploidy screening by QF-PCR amplification of short tandem repeats (STRs) on chromosomes X, Y, 21, 13, and 18 was carried out within 24 h of collection. Owing to the sampling procedure, the eventual presence of contamination between fetuses was also evaluated in every case. RESULTS: Normal and aneuploid fetuses were readily identified by QF-PCR. Fetal reduction was made available, for trisomic fetuses, without further waiting for completion of fetal karyotyping. In twin gestations, the ultrasound examination of chorionicity was always in agreement with the molecular assessment of zygosity. Contamination between fetuses due to the sampling procedure with a single puncture was never observed. CONCLUSION: Rapid prenatal diagnosis of aneuploidies by QF-PCR is a sensitive, efficient, and reliable assay. When applied in multiple pregnancies, it has the added value of allowing the assessment of zygosity in all cases, independently of chorionicity and fetal sex.

Targeted expression of human serum albumin to potato tubers.

Complementary DNA expression of mature human serum albumin was engineered into potato plants under the transcriptional control of patatin B33 promoter and potato proteinase inhibitor II terminator. Protein secretion was achieved by using the signal sequence from potato proteinase inhibitor II. Recombinant albumin accumulated up to 0.2% of total soluble tuber protein in single transformant lines, regardless of the potato cultivar used. Electrophoretic mobility and N-terminal amino acid sequence analysis of partially purified recombinant albumin confirmed proper processing of an immune responsive recombinant albumin, and revealed that the proteinase inhibitor II signal sequence was correctly removed. No further optimisation of these yields was obtained by HSA expression in patatin antisense plants (line Pas58). Subcellular localisation showed that recombinant protein was successfully targeted to the apoplast. Potato tubers may be used, by applying this technology, to produce other heterologous proteins of interest in the biopharmaceutical industry.