This gene sequencing panel includes both sequencing and deletion/duplication (CNV) analysis for all coding regions of the included genes (unless otherwise noted). All analysis is performed utilizing next-generation sequencing (NGS) technology. All variants are classified according to American College of Genetics and Genomics (ACMG) guidelines.

Spastic paraplegia refers to a group of neurological disorders characterized by progressive weakness and spasticity in the muscles of the lower extremities. This condition primarily affects the ability to walk and can result in varying degrees of disability. Individuals with spastic paraplegia experience progressive weakness in the muscles of the lower limbs, leading to difficulties with walking, climbing stairs, and maintaining balance. The weakness typically begins insidiously and worsens over time. Spasticity refers to increased muscle tone and stiffness, which can cause muscles to become rigid and resistant to movement, leading to difficulty with voluntary movements. Spastic paraplegia often results in changes to the individual's gait, including a stiff-legged or "scissored" gait, where the legs cross over each other with each step. Walking may become slower, more laborious, and less coordinated as the condition progresses. While spasticity and weakness primarily affect the muscles of the lower limbs, individuals with spastic paraplegia may also experience involvement of the upper limbs, although to a lesser extent. Some individuals with spastic paraplegia may experience sensory changes, such as numbness, tingling, or loss of sensation in the lower extremities. These sensory changes may be secondary to compression of nerves or other neurological abnormalities. Spastic paraplegia can also lead to urinary and bowel dysfunction, including urinary urgency, frequency, and incontinence, as well as constipation and difficulty with bowel movements. The progression of spastic paraplegia can vary widely among affected individuals and between different genetic subtypes. While some individuals may experience relatively slow progression and maintain functional independence for many years, others may experience more rapid deterioration and significant disability.

ALS2, ATL1, DDHD1, HSPD1, NT5C2, SLC16A2, TFG, AP4S1, CCT5, FA2H, MAG, REEP2, SPG21, ZFYVE26, AP4B1, B4GALNT1, ENTPD1, KIF1A, PLP1, SPART, VAMP1, ALDH18A1, ARL6IP1, CYP7B1, GJC2, NIPA1, SACS, TECPR2, AP4M1, C19orf12, ERLIN2, KIF5A, REEP1, SPG11, WASHC5, AMPD2, ATP13A2, DDHD2, IBA57, PGAP1, SLC33A1, TTR, AP5Z1, CYP2U1, GBA2, MTRFR, RTN2, SPG7, AP4E1, BSCL2, ERLIN1, KIF1C, PNPLA6, SPAST, VPS37A

Sequencing is performed on genomic DNA using an Agilent targeted sequence capture method to enrich for the exome. Direct sequencing of the amplified captured regions was performed using 2X150bp reads on Illumina next generation sequencing (NGS) systems. A base is considered to have sufficient coverage at 20X and an exon is considered fully covered if all coding bases plus three nucleotides of flanking sequence on either side are covered at 20X or more. A list of these regions, if any, is available upon request. Alignment to the human reference genome (GRCh37) is performed and annotated variants are identified in the targeted region. Variants reviewed have a minimum coverage of 8X and an alternate allele frequency of 20% or higher. Indel and single nucleotide variants (SNVs) may be confirmed by Sanger sequence analysis before reporting at director discretion. This assay cannot detect variants in regions of the exome that are not covered, such as deep intronic, promoter and enhancer regions, areas containing large numbers of tandem repeats, and variants in mitochondrial DNA. Copy number variation (CNV) analysis detects deletions and duplications; in some instances, due to the size of the exons, sequence complexity, or other factors, not all CNVs may be analyzed or may be difficult to detect. When reported, copy number variant size is approximate. Actual breakpoint locations may lie outside of the targeted regions. CNV analysis will not detect tandem repeats, balanced alterations (reciprocal translocations, Robertsonian translocations, inversions, and balanced insertions), methylation abnormalities, triploidy, and genomic imbalances in segmentally duplicated regions. This assay is not designed to detect mosaicism; possible cases of mosaicism may be investigated at the discretion of the laboratory director. Primary data analysis is performed using Illumina bcl2fastq converter v2.19. Secondary analysis is performed using Illumina DRAGEN Bio-IT Platform v.3.10.8. Tertiary data analysis is performed using SnpEff v5.0 and Revvity Omics’ internal ODIN v.1.01 software. CNV and absence of heterozygosity are assessed using BioDiscovery’s NxClinical v6.1 software.