Event Title

AGG Interruptions: Improving Fragile X Screening at NxGen MDx

Location

Hager-Lubbers Exhibition Hall

Start Date

15-4-2019 3:30 PM

Description

PURPOSE: Mutations in the fragile X mental retardation 1 (FMR1) gene cause fragile X syndrome when the number of trinucleotide CGG repeats is 200 or greater. Normal genotypes for FMR1 contain 5 to 44 repeats and 0 to 3 AGG interruptions, which are AGG sequences within the FMR1 CGG repeat region to stabilize the sequence and anchor the polymerase to the correct sequence to increase the precision of replication. CHALLENGE: NxGen MDx used a screening procedure to determine which patients were normal or at risk of expansion. This was only successful up to eighty repeats and required several samples to be sent to an external company for further characterization. EXPERIENCE: Using PCR-based fragment analysis, a fragile X screening methodology was developed for intermediate mutations and another to characterize full mutations. A repeat chimeric primer was utilized to determine repeat number and AGG interruption status within the FMR1 gene. OUTCOME: Each protocol was optimized for primer concentrations, cycling parameters, and master mix composition. Modifications to the capillary electrophoresis program were also essential to characterize a full mutation. The improved fragile X methodology adapted by NxGen MDx eliminated the need to send samples out externally and minimized sample reruns. IMPACT: The opportunity to intern with NxGen MDx validated experiences from classes using molecular techniques. It was rewarding to contribute to a substantial project for a small, local biotechnology company. Shortly after my internship concluded, I was hired as clinical molecular technician to continue working at NxGen MDx.

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Apr 15th, 3:30 PM

AGG Interruptions: Improving Fragile X Screening at NxGen MDx

Hager-Lubbers Exhibition Hall

PURPOSE: Mutations in the fragile X mental retardation 1 (FMR1) gene cause fragile X syndrome when the number of trinucleotide CGG repeats is 200 or greater. Normal genotypes for FMR1 contain 5 to 44 repeats and 0 to 3 AGG interruptions, which are AGG sequences within the FMR1 CGG repeat region to stabilize the sequence and anchor the polymerase to the correct sequence to increase the precision of replication. CHALLENGE: NxGen MDx used a screening procedure to determine which patients were normal or at risk of expansion. This was only successful up to eighty repeats and required several samples to be sent to an external company for further characterization. EXPERIENCE: Using PCR-based fragment analysis, a fragile X screening methodology was developed for intermediate mutations and another to characterize full mutations. A repeat chimeric primer was utilized to determine repeat number and AGG interruption status within the FMR1 gene. OUTCOME: Each protocol was optimized for primer concentrations, cycling parameters, and master mix composition. Modifications to the capillary electrophoresis program were also essential to characterize a full mutation. The improved fragile X methodology adapted by NxGen MDx eliminated the need to send samples out externally and minimized sample reruns. IMPACT: The opportunity to intern with NxGen MDx validated experiences from classes using molecular techniques. It was rewarding to contribute to a substantial project for a small, local biotechnology company. Shortly after my internship concluded, I was hired as clinical molecular technician to continue working at NxGen MDx.