High-quality read-based phasing of cystic fibrosis cohort informs genetic understanding of disease modification

Mastromatteo, Scott; Chen, Angela; Gong, Jiafen; Fan, Lin; Thiruvahindrapuram, Bhooma; Sung, Wilson W L; Whitney, J. Andrew; Wang, Zhuozhi; Patel, Rohan; Keenan, Katherine; Halevy, Ayelet; Panjwani, Naim; Avolio, Julie; Wang, Cheng; Côté-Maurais, Guillaume; Bégin, Stéphanie; Adam, Damien; Brochiero, Emmanuelle; Bjornson, Candice; Chilvers, Mark; Price, April; Parkins, Michael D.; Wylick, Richard van; Mateos-Corral, Dimas; Hughes, Daniel; Smith, Mary Jane; Morrison, Nancy; Tullis, Elizabeth; Stephenson, Anne L.; Wilcox, Pearce; Quon, Bradley S.; Leung, Winnie; Solomon, Melinda; Sun, Lei; Ratjen, Félix; Strug, Lisa J.

doi:10.1016/j.xhgg.2022.100156

Cited by 2 publications

(2 citation statements)

References 55 publications

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“…Long-range information embedded into short reads is essential for interpreting genomic data for many different applications 17,[47][48][49][50][51][52][53][54] . Single-tube TELL-Seq is an instrument-free linked-read method characterized by its versatility and simplicity 24 .…”

Section: Discussionmentioning

confidence: 99%

Targeted Phasing of 2-200 Kilobase DNA Fragments with a Short-Read Sequencer and a Single-Tube Linked-Read Library Method

Mikhaylova¹,

Rzepka²,

Kawamura³

et al. 2023

Preprint

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In the human genome, heterozygous sites are genomic positions with different alleles inherited from each parent. On average, there is a heterozygous site every 1-2 kilobases (kb). Resolving whether two alleles in neighboring heterozygous positions are physically linked—that is, phased—is possible with a short-read sequencer if the sequencing library captures long-range information. TELL-Seq is a library preparation method based on millions of barcoded micro-sized beads that enables instrument-free phasing of a whole human genome in a single PCR tube. TELL-Seq incorporates a unique molecular identifier (barcode) to the short reads generated from the same high-molecular-weight (HMW) DNA fragment (known as ‘linked-reads’). However, genome-scale TELL-Seq is not cost-effective for applications focusing on a single locus or a few loci. Here, we present an optimized TELL-Seq protocol that enables the cost-effective phasing of enriched loci (targets) of varying sizes, purity levels, and heterozygosity. Targeted TELL-Seq maximizes linked-read efficiency and library yield while minimizing input requirements, fragment collisions on microbeads, and sequencing burden. To validate the targeted protocol, we phased seven 180-200 kb loci enriched by CRISPR/Cas9-mediated excision coupled with pulse-field electrophoresis, four 20 kb loci enriched by CRISPR/Cas9-mediated protection from exonuclease digestion, and six 2-13 kb loci amplified by PCR. The selected targets have clinical and research relevance (BRCA1, BRCA2, MLH1, MSH2, MSH6, APC, PMS2, SCN5A-SCN10A, andPKI3CA). These analyses reveal that targeted TELL-Seq provides a reliable way of phasing allelic variants within targets (2-200 kb in length) with the low cost and high accuracy of short-read sequencing.

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Section: Discussionmentioning

confidence: 99%

Targeted Phasing of 2-200 Kilobase DNA Fragments with a Short-Read Sequencer and a Single-Tube Linked-Read Library Method

Mikhaylova¹,

Rzepka²,

Kawamura³

et al. 2023

Preprint

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show abstract

“…It should however be emphasized that c.-204A>C may not be the sole functional variant underlying the increased risk of the c.-408T>C-tagged haplotype. This risk haplotype has been recently shown to contain the PRSS3P2 and TRY7 pseudogenes whereas the alternative haplotype has lost the two pseudogenes [50,51]; this alternative haplotype was reported to be associated with a protective effect against CP [52]. Interestingly, the risk haplotype appears to contain still functional PRSS3P2/TRY7 pseudogene enhancers that serve to upregulate pancreatic PRSS2 expression [53], which is consistent with the increasingly appreciated role of PRSS2 in J o u r n a l P r e -p r o o f pancreatitis (see Masson et al [52] and references therein).…”

Section: Common Variantsmentioning

confidence: 99%

Classification of PRSS1 variants responsible for chronic pancreatitis: An expert perspective from the Franco-Chinese GREPAN Study Group

Masson¹,

Zou²,

Pu³

et al. 2023

Pancreatology

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High-quality read-based phasing of cystic fibrosis cohort informs genetic understanding of disease modification

Cited by 2 publications

References 55 publications

Targeted Phasing of 2-200 Kilobase DNA Fragments with a Short-Read Sequencer and a Single-Tube Linked-Read Library Method

Targeted Phasing of 2-200 Kilobase DNA Fragments with a Short-Read Sequencer and a Single-Tube Linked-Read Library Method

Classification of PRSS1 variants responsible for chronic pancreatitis: An expert perspective from the Franco-Chinese GREPAN Study Group

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