Population Scale HLA-Typing and Uses Thereof

Abstract

The present invention provides a portable system for real-time population-scale HLA genotyping and/or allelotyping in a field environment and methods of such population-scale HLA genotyping. The individual components of the system are portable to and operable within a field environment thereby providing high throughput with real-time geno- or allelotyping. Also provided are HLA gene-specific primers and HLA allele-specific or single nucleotide polymorphism-specific hybridization probes. In addition the present invention provides a microarray comprising the hybridization probes. Further provided is a kit comprising the HLA gene-specific primers and the microarray.

What is claimed is: 
     
         1 . Hybridization probes for detecting single nucleotide polymorphisms (SNPs) in an HLA gene, wherein said hybridization probes are selected from the group consisting of consisting of SEQ ID NOS: 49-99, 101-158, 160-161, 163-228, 230-239, 241, 243-283, and 285-289. 
     
     
         2 . A microarray device for allelotyping an HLA gene, comprising:
 a substrate having a cationic surface; and   a monolayer comprising one or more of the hybridization probes of  claim 1  adsorbed thereto.   
     
     
         3 . The microarray device of  claim 2 , wherein the cationic surface comprises an aminosilane, a quanidinium, tin oxide, aluminum oxide or zirconium oxide or other equivalently charged moiety. 
     
     
         4 . The microarray device of  claim 2 , wherein the substrate is glass, plastic or metal. 
     
     
         5 . The microarray device of  claim 2 , further comprising one or more of:
 an oligo-thymidine co-absorbed with the hybridization probes;   a fluorescent dye linked to the oligo-thymidine; or   a capping agent.   
     
     
         6 . The microarray device of  claim 2 , wherein the oligo-thymidine has about 20 to about 40 thymidines. 
     
     
         7 . A kit for population-scale HLA genotyping, comprising:
 gene-specific primers for amplifying an HLA gene having the sequences shown in SEQ ID NOS: 14-47; and   a microarray device for allelotyping the HLA gene, comprising:
 a substrate having a cationic surface; and 
 a monolayer comprising one or more hybridization probes from a group consisting of the sequences shown in SEQ ID NOS: 48-289 adsorbed thereto. 
   
     
     
         8 . The kit of  claim 7 , wherein the cationic surface of the microarray device comprises an aminosilane, a quanidinium, tin oxide, aluminum oxide or zirconium oxide or other equivalently charged moiety. 
     
     
         9 . The kit of  claim 7 , wherein the substrate is glass, plastic or metal. 
     
     
         10 . The kit of  claim 7 , further comprising one or more of:
 an oligo-thymidine comprising 20 to about 40 thymidine co-absorbed with the hybridization probes;   a fluorescent dye linked to the oligo-thymidine; or   a capping agent.   
     
     
         11 . The kit of  claim 7 , further comprising:
 buffers and polymerases for a PCR reaction or a fluorescent dye or a combination thereof.   
     
     
         12 . A system for real-time high throughput population-scale HLA allelotyping in a field environment, comprising:
 the kit of  claim 11 ;   means for collecting and purifying DNA samples from individuals comprising a population; and   means for assigning an HLA allelotype to each individual HLA gene of interest; wherein individual means and devices comprising said system are portable and operable in real time within the field environment.   
     
     
         13 . The system of  claim 12 , wherein the means for collecting DNA samples comprises a container suitable to receive a buccal wash sample, a buccal swab sample or a blood sample collected from the individuals. 
     
     
         14 . The system of  claim 12 , wherein said means for assigning an HLA-allelotype to each individual comprises:
 an imaging device adapted to detect hybridization patterns formed on the microarray device after hybridization of the target to the hybridization probes adsorbed thereto; and   pattern recognition software comprising a set of algorithms adapted to recognize the imaged hybridization patterns as HLA allelotypes.   
     
     
         15 . The system of  claim 12 , wherein real time high throughput genotyping is about 200 to about 300 HLA allelotypes per hour per system operated. 
     
     
         16 . A method for real time population-scale HLA allelotyping in a field environment, comprising the steps of:
 collecting DNA from one or more members of the population;   purifying the DNA for analysis;   generating a target amplicon from an HLA gene of interest comprising the DNA using gene specific primers;   contacting the hybridization probes comprising the microarray of the kit of  claim 11  with the target; and   imaging the hybridization pattern formed after the contact wherein each HLA allelotype has a pattern associated therewith.   
     
     
         21 . The method of  claim 16 , further comprising storing the collected DNA. 
     
     
         22 . The method of  claim 16 , wherein the DNA is collected from blood, with a buccal wash or with a buccal swab. 
     
     
         23 . The method of  claim 16 , further comprising one or both of:
 assessing a risk of infection by a biological agent or weapon for each individual based on the assigned allelotype; or   assessing a response to a particular vaccine against the biological agent or weapon by each individual; or   
     
     
         25 . The method of  claim 16 , wherein the allelotype assigned to each individual of the population comprises a means of identification thereof.