Real-Time Quantitative PCR for Detection Cell Free Fetal DNA

Basically, diagnosis inherited diseases in gestation is very important. But amplication of the progress in early stage of pregnance is the serious and great aim for rapid application of risk assesment for both and fetus. Fetal genetic tissues collected through techniques such as amniocentesis and chorionic villus sampling (CVS) for prenatal diagnosis of fetal genetic diseases. These procedures are associated with a risk of fetal loss (1%) and two week needed for cultivation (Chiu et al., 2010). Rapid methods for prenatal diagnosis of fetal chromosomal aneuploidies have been developed. This method is multicolor fluorescence in situ hybridization (FISH). FISH is very reliable but requires intact cells and it can only be used on fresh samples (Klinger et al.,1992) The other method is quantitative fluorescent polymerase chain reaction PCR anaylsis (real-time PCR) (Lo et al., 1997). Real-time PCR is a powerful tool for quantifying gene expression combining both high sensitivity and specificity with efficient signal detection (Kubista, 2008). A major advantage of real-time PCR is that it can be used to determine the amount of initial temlates (Heidi et al., 1996). In addition, by making use of a closed whereby the samples are analyzed directly in PCR reaction vessel during the amplification. Furthermore, the assay is less prone contamination (Zimmerman, 2006). Real-time PCR has found wide spread applicability in the analysis of gene expression measurement and cell-free nucleic acids in body fluids. The assay is readily amenable to automation, and by making use of the current real-time PCR 96-384 well formats. The discovery of cff DNA in maternal plasma in 1997 has opened non-invasive prenatal diagnosis (NIPD). This research area is a rapidly developing and dynamic field. NIPD using cell-free fetal DNA (and RNA) is likely to become increasingly available within the next few years. There have been a lot of reported applications, including fetal rhesus-D genotyping (Bombard et al.,2011), fetal sexing for X-linked disorders (Costa et al., 2002), paternally inherited genetic diseases, and pregnancy-associated conditions such as preeclampsia (Zhong et al., 2001). Diagnosing by using free DNA a variety of limitations due to some of the features of this resource. CffDNA analyses would be applicable only when the involved target sequence is in the fetus but absent from mother, such as paternally