MICROARRAY MEETING 2002: NEW DEVELOPMENTS IN MUTATION DETECTION AND GENE EXPRESSION
Segrate, MI (Italy), April 12, 2002 

Minerva Biotecnologica 2002 December;14(3-4):237-40

Copyright © 2003 EDIZIONI MINERVA MEDICA

language: English

Genetic diseases. Development of a rapid protocol for mutation detection by microarray analysis

Carella M. ^{1, 2}, Stanziale P. ², Gasparini P. ^{1, 3}

¹ TIGEM (Telethon Institute of Genetics and Medicine), Naples; ² Service of Medical Genetics, “CSS” Hospital-IRCCS, San Giovanni Rotondo, Foggia; ³ Department of General Pathology, Medical Genetics, Second University of Naples, Naples

Sev­er­al genet­ic dis­eas­es are char­ac­ter­ized by ­genes ­with one or few ­very fre­quent muta­tions. ­Here we ­report the devel­op­ment of a new pro­to­col ­based on ­arrayed prim­er exten­sion (­APEX) tech­nique for the detec­tion of ­very com­mon ­alleles. In ­this new pro­to­col, ­named ­arrayed ampli­con prim­er exten­sion (AAP­EX), for the ­first ­time ­instead of link­ing ami­no-­mod­i­fied oli­go­nu­cleo­tides on the ­glass sur­face, we ­have ­attached on the ­same sur­face PCR prod­ucts cor­re­spond­ing to dif­fer­ent ­patients and car­riers. ­These prod­ucts ­have ­been ­obtained ­using 1 ami­no­-mod­i­fied oli­go out of the 2 PCR prim­ers. ­This pro­to­col has ­been devel­oped for the detec­tion of 1 of the 2 ­most com­mon ­alleles (H63D) of HFE ­gene, ­which is mutat­ed in hered­i­tary hemo­chro­mat­o­sis, one of the ­most com­mon reces­sive dis­eas­es in Cau­ca­sians. A ­series of experi­ments per­formed ­with ­this pro­to­col ­allowed us to dem­on­strate ­that it can be eas­i­ly ­applied to the simul­ta­ne­ous screen­ing of sev­er­al indi­vid­u­als. Our ­results clear­ly dem­on­strate the fea­sibil­ity, reli­abil­ity and spec­i­fic­ity of ­this pro­to­col for a muta­tion­al screen­ing of a ­large ­series of indi­vid­u­als.