scholarly journals Single molecule sequencing of THCA synthase reveals copy number variation in modern drug-type Cannabis sativa L.

2015 ◽  
Author(s):  
Kevin J McKernan ◽  
Yvonne Helbert ◽  
Vasisht Tadigotla ◽  
Stephen McLaughlin ◽  
Jessica Spangler ◽  
...  
Keyword(s):  
Copy Number Variation ◽  
Single Molecule ◽  
Copy Number ◽  
Cannabis Sativa ◽  
Early Stage ◽  
Single Molecule Sequencing ◽  
Modern Drug ◽  
Number Variation ◽  
Complex Picture ◽  
Genetically Determined

Cannabinoid expression is an important genetically determined feature of cannabis that presents clinical and legal implications for patients seeking cannabinoid specific therapies like Cannabidiol (CBD). Cannabinoid, terpenoid, and flavonoid marker assisted selection can accelerate breeding efforts by offering genetic tools to select for desired traits at an early stage in growth. To this end, multiple models for chemotype inheritance have been described suggesting a complex picture for chemical phenotype determination. Here we explore the potential role of copy number variation of THCA Synthase using phased single molecule sequencing and demonstrate that copy number and sequence variation of this gene is common and suggests a more nuanced view of chemotype prediction.

Sequence and annotation of 42 cannabis genomes reveals extensive copy number variation in cannabinoid synthesis and pathogen resistance genes

2020 ◽  
Author(s):  
Kevin J. McKernan ◽  
Yvonne Helbert ◽  
Liam T. Kane ◽  
Heather Ebling ◽  
Lei Zhang ◽  
...  
Keyword(s):  
Copy Number Variation ◽  
Single Molecule ◽  
Resistance Genes ◽  
Copy Number ◽  
Cannabis Sativa ◽  
Pathogen Resistance ◽  
Single Molecule Sequencing ◽  
Model Predictions ◽  
Long Read ◽  
Number Variation

AbstractCannabis is a diverse and polymorphic species. To better understand cannabinoid synthesis inheritance and its impact on pathogen resistance, we shotgun sequenced and assembled a Cannabis trio (sibling pair and their offspring) utilizing long read single molecule sequencing. This resulted in the most contiguous Cannabis sativa assemblies to date. These reference assemblies were further annotated with full-length male and female mRNA sequencing (Iso-Seq) to help inform isoform complexity, gene model predictions and identification of the Y chromosome. To further annotate the genetic diversity in the species, 40 male, female, and monoecious cannabis and hemp varietals were evaluated for copy number variation (CNV) and RNA expression. This identified multiple CNVs governing cannabinoid expression and 82 genes associated with resistance to Golovinomyces chicoracearum, the causal agent of powdery mildew in cannabis. Results indicated that breeding for plants with low tetrahydrocannabinolic acid (THCA) concentrations may result in deletion of pathogen resistance genes. Low THCA cultivars also have a polymorphism every 51 bases while dispensary grade high THCA cannabis exhibited a variant every 73 bases. A refined genetic map of the variation in cannabis can guide more stable and directed breeding efforts for desired chemotypes and pathogen-resistant cultivars. Sequence and annotation of 42 cannabis genomes reveals extensive copy number variation in cannabinoid synthesis and pathogen resistance genes

BRCA Testing by Single-Molecule Molecular Inversion Probes

2017 ◽  
Vol 63 (2) ◽  
pp. 503-512 ◽  
Author(s):  
Kornelia Neveling ◽  
Arjen R Mensenkamp ◽  
Ronny Derks ◽  
Michael Kwint ◽  
Hicham Ouchene ◽  
...  
Keyword(s):  
Genetic Testing ◽  
Copy Number Variation ◽  
Single Molecule ◽  
Copy Number ◽  
Single Gene ◽  
Work Flow ◽  
Analytical Sensitivity ◽  
Brca1 And Brca2 ◽  
Number Variation ◽  
Molecular Inversion Probes

Abstract BACKGROUND Despite advances in next generation DNA sequencing (NGS), NGS-based single gene tests for diagnostic purposes require improvements in terms of completeness, quality, speed, and cost. Single-molecule molecular inversion probes (smMIPs) are a technology with unrealized potential in the area of clinical genetic testing. In this proof-of-concept study, we selected 2 frequently requested gene tests, those for the breast cancer genes BRCA1 and BRCA2, and developed an automated work flow based on smMIPs. METHODS The BRCA1 and BRCA2 smMIPs were validated using 166 human genomic DNA samples with known variant status. A generic automated work flow was built to perform smMIP-based enrichment and sequencing for BRCA1, BRCA2, and the checkpoint kinase 2 (CHEK2) c.1100del variant. RESULTS Pathogenic and benign variants were analyzed in a subset of 152 previously BRCA-genotyped samples, yielding an analytical sensitivity and specificity of 100%. Following automation, blind analysis of 65 in-house samples and 267 Norwegian samples correctly identified all true-positive variants (>3000), with no false positives. Consequent to process optimization, turnaround times were reduced by 60% to currently 10–15 days. Copy number variants were detected with an analytical sensitivity of 100% and an analytical specificity of 88%. CONCLUSIONS smMIP-based genetic testing enables automated and reliable analysis of the coding sequences of BRCA1 and BRCA2. The use of single-molecule tags, double-tiled targeted enrichment, and capturing and sequencing in duplo, in combination with automated library preparation and data analysis, results in a robust process and reduces routine turnaround times. Furthermore, smMIP-based copy number variation analysis could make independent copy number variation tools like multiplex ligation-dependent probes amplification dispensable.

Identification of Novel Germline and Tumor-Specific Nucleotide Variants and Copy Number Variation in Clival Chordomas by Exome Sequencing

2015 ◽  
Vol 76 (S 01) ◽  
Author(s):  
Georgios Zenonos ◽  
Peter Howard ◽  
Maureen Lyons-Weiler ◽  
Wang Eric ◽  
William LaFambroise ◽  
...  
Keyword(s):  
Copy Number Variation ◽  
Exome Sequencing ◽  
Copy Number ◽  
Number Variation ◽  
Specific Nucleotide

The impact of paralog genes: detection of copy number variation in spinal muscle atrophy patients

BIOCELL ◽  
2018 ◽  
Vol 42 (3) ◽  
pp. 87-91 ◽  
Author(s):  
Sergio LAURITO ◽  
Juan A. CUETO ◽  
Jimena PEREZ ◽  
Mar韆 ROQU�
Keyword(s):  
Copy Number Variation ◽  
Muscle Atrophy ◽  
Copy Number ◽  
Number Variation ◽  
The Impact
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