Microarray-based assay integrated with particles for analyzing molecular interactions
A microarray-based assay is provided, which is used for analyzing molecular interactions, including polynucleotides, polypeptides, antibodies, small molecule compounds, peptides and carbohydrates. Such method comprises coupling a target molecule to a particle and then binding to a probe molecule on microarray. In particular, multiplexed genetic analysis of nucleic acid fragments can be implemented. Specific genes, single nucleotide polymorphisms or gene mutations, such as deletions, insertions, and indels, can be identified. Coupled with microarray, the particles, themselves or further modified, facilitate the detection of results with non-expensive devices or even naked eyes. This technology enables the detection and interpretation of molecular interactions in an efficient and cost effective way.
1. A method for detecting a target molecule using a microarray, which method comprises:
a) coupling the target molecule to a particle of a diameter from about 0.1 μm to about 1 mm;
b) allowing the target molecule to interact with a probe molecule immobilized on a spot on the microarray, wherein the spot has a diameter from about 0.01 mm to about 5 mm; and
c) detecting the interaction between the target molecule and the probe molecule by visualizing the particle on the spot under a visible light and/or using fluorescence scanning,
wherein the target molecule is selected from the group consisting of a polynucleotide, a polypeptide, an antibody, a small molecule compound, a peptide and a carbohydrate, and
wherein the microarray comprises a universal Tag array comprising at least two of the Tag sequences as set forth in Table 1.
2. A method for detecting a target molecule using a microarray, which method comprises:
a) coupling a double stranded target molecule to a particle of a diameter from about 0.1 μm to about 1 mm;
b) recovering a single stranded target molecule coupled to the particle from the double stranded target molecule;
c) allowing the single stranded target molecule to interact with a probe molecule immobilized on a spot on the microarray, wherein the spot has a diameter from about 0.01 mm to about 5 mm; and
d) detecting the interaction between the target molecule and the probe molecule by visualizing the particle on the spot under a visible light and/or using fluorescence scanning,
wherein the target molecule is a polynucleotide, and
wherein the microarray comprises a universal Tag array comprising at least two of the Tag sequences as set forth in Table 1.
3. The method according to claim 1 or 2 , wherein the particle and/or the target molecule is modified with a functional group selected from the group consisting of a chemical group, a polynucleotide, a polypeptide, an antibody, a small molecule compound, a peptide and a carbohydrate.
4. The method of claim 3 , wherein both the particle and the target molecule are modified and the target molecule is coupled to the particle through an interaction between the modification of the target molecule and the functional group on the particle.
5. The method of claim 3 , wherein the interaction between the target molecule and the probe molecule is a non-covalent interaction.
6. The method of claim 5 , wherein the efficiency and/or efficacy of the interaction is enhanced by a magnetic force, a dielectrophoretic force, a mechanical force, or a combination thereof.
7. The method of claim 2 , wherein the target polynucleotide is subject to an in vitro manipulation, wherein the in vitro manipulation is selected from the group consisting of laser treatment, ultrasonication, heating, microwave treatment, piezoelectricity treatment, electrophoresis, dielectrophoresis, solid phase adhesion, filtration, fluidic stress treatment, enzymatic digestion, PCR amplification, reverse-transcription, reverse-transcription PCR amplification, allele-specific PCR (ASPCR), single-base extension (SBE), allele specific primer extension (ASPE), restriction enzyme digestion, strand displacement amplification (SDA), transcription mediated amplification (TMA), ligase chain reaction (LCR), nucleic acid sequence based amplification (NASBA), primer extension, rolling circle amplification (RCA), self sustained sequence replication (3SR), the use of Q Beta replicase, nick translation, and loop-mediated isothermal amplification (LAMP).
8. The method of claim 2 , wherein the target polynucleotide comprises a genetic information.
9. The method of claim 8 , wherein the genetic information associated with the target polynucleotide is a mutation selected from the group consisting of a substitution, an insertion, a deletion and an indel.
10. The method of claim 9 , wherein the mutation is a single nucleotide polymorphism (SNP).
11. The method according to claim 8 , wherein the genetic information is associated with: a disease caused by an infectious or pathogenic agent selected from the group consisting of a fungus, a bacterium, a mycoplasma , a rickettsia , a chlamydia , a virus and a protozoa; a sexually transmitted disease; cancer; cerebrovascular disease; heart disease; respiratory disease; coronary heart disease; diabetes; hypertension; Alzheimer's disease; neurodegenerative disease; chronic obstructive pulmonary disease; autoimmune disease; cystic fibrosis; spinal muscular atrophy; beta thalassemia; phenylalanine hydroxylase deficiency; Duchenne muscular dystrophy; or hereditary hearing loss.
12. The method of claim 11 , wherein the genetic information is associated with hereditary hearing loss.
13. The method of claim 1 , wherein the diameter of the particle is from about 0.1 μm to about 10 μm.
14. The method of claim 2 , wherein the diameter of the particle is from about 0.1 μm to about 10 μm.
15. The method of claim 1 , wherein the diameter of the spot on the microarray is from about 0.03 mm to about 5 mm.
16. The method of claim 2 , wherein the diameter of the spot on the microarray is from about 0.03 mm to about 5 mm.
17. The method of claim 3 , wherein the interaction between the target molecule and the probe molecule is a reversible covalent or irreversible covalent interaction.
18. The method of claim 1 or 2 , wherein the microarray comprises all of the Tag sequences set forth in Table 1.
19. The method of claim 7 , wherein the allelic-specific PCR (ASPCR) comprises using a set of primers for ASPCR amplification of a genetic information, wherein the set of primers comprises two allele-specific primers and a common primer as set forth in Table 2.