The objectives of this study is to check what gene is suitable for creation of a standard,to establish real time PCR as a high throughput technique for copy number work and to optimize / prepare a good standard for absolute quantification analysis for this work.
We designed two strategies for the optimization of qRT-PCR. The first one is through Southern blot copy number correlation with qRT-PCR. The second is through the creation of standard using a plasmid carrying the construct that was used in the plant.
In lieu of the first objective to find a gene suitable for doing copy number work we encountered problem with NOS gene.
We did the serial dilution of the samples following the computation by Applied Biosystem.
http://mponscientic.blogspot.com/2010/05/copy-number-determination-optimisation.html