In this study, a novel method for the direct detection of GMP without amplified by the general method of PCR is firstly
presented and proved by experiments. In our method, fluorescence correlation spectroscopy, cleaving nucleic acid by
restriction endonuclease and two nucleic acid probe hybridization techniques are combined to distinguish the
caulifiower mosaic virus (CaMV) 35S promoter and determine whether samples contain genetically modified
components. The detection principle is as follows: firstly two restriction endonucleases FOKI and BsrDlare used to
cleave the genomic DNA and the 169bp fragments of CaMV 35S promoter are retrieved; secondly, two nucleic acid
probes labeled by Rhodamine Green and y5 dyes respectively hybridize with cleaved 169bp fragments of CaMV 35S
promoter; thirdly, the hybridization products simultaneously with two dye-labeled probes are detected by fluorescence
cross-correlation spectroscopy and GMP is distinguished. As the detection and analysis by FCS can be performed at the
level of single molecule, there is no need for any type of amplification. Genetically modified tobaccos are measured by
this method. The results indicate this method can detect CaMV 35S promoter of GMP exactly and the sensitivity can be
down to 3.47X10-10M. Because no any type of amplification is involved, this method can avoid the non-specffic
amplification and false-positive problems of PCR, Due to its high-sensitivity, simplicity, reliability and little need for
sample amounts, this method promises to be a highly effective detection method for GMP.
The concentration dependence of the diffusion coefficient of particles suspended in solution depends primarily on the occupied volume fraction and on repulsive and attractive forces. This dependency is expressed by the interaction parameter λ. In the present work we have measured the diffusion coefficient of Bovine Serum Albumin at different ionic strength and determined the interaction parameter. The results indicate: the value of λ is positive at low ionic strength and the interaction between proteins is repulsive; however, with increasing ionic strength the value of λ becomes negative and the interaction is attractive, and when ionic strength is bigger than 0.50M aggregation occurs. The dependence of the interaction on ionic strength is interpreted using DLVO theory for interactions of two hard spheres: with increasing ionic strength, the repulsive electrostatic interaction is screened and Van der Waals forces become dominant. According to the correlation of λ with ionic strength, the protein parameters are regressed: the protein net charge ZP= -9.0e, Hamaker constant HA= 2.8kBT. This work indicates the technique of dynamic light scattering can be used effectively to study protein molecular interactions.