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Summary From a single transgenic line harboring fiveTnt1transposon insertions, we generated a near‐saturated insertion population inMedicago truncatula. Using thermal asymmetric interlaced‐polymerase chain reaction followed by sequencing, we recovered 388 888 flanking sequence tags (FSTs) from 21 741 insertion lines in this population.FSTrecovery from 14Tnt1lines using the whole‐genome sequencing (WGS) and/orTnt1‐capture sequencing approaches suggests an average of 80 insertions per line, which is more than the previous estimation of 25 insertions. Analysis of the distribution pattern and preference ofTnt1insertions showed thatTnt1is overall randomly distributed throughout theM. truncatulagenome. At the chromosomal level,Tnt1insertions occurred on both arms of all chromosomes, with insertion frequency negatively correlated with theGCcontent. Based on 174 546 filteredFSTs that show exact insertion locations in theM. truncatulagenome version 4.0 (Mt4.0), 0.44Tnt1insertions occurred per kb, and 19 583 genes containedTnt1with an average of 3.43 insertions per gene. Pathway and gene ontology analyses revealed thatTnt1‐inserted genes are significantly enriched in processes associated with ‘stress’, ‘transport’, ‘signaling’ and ‘stimulus response’. Surprisingly, gene groups with higher methylation frequency were more frequently targeted for insertion. Analysis of 19 583Tnt1‐inserted genes revealed that 59% (1265) of 2144 transcription factors, 63% (765) of 1216 receptor kinases and 56% (343) of 616 nucleotide‐binding site‐leucine‐rich repeat genes harbored at least oneTnt1insertion, compared with the overall 38% ofTnt1‐inserted genes out of 50 894 annotated genes in the genome.
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Leveraging whole‐genome sequencing to estimate telomere length in plants
Abstract Changes in telomere length are increasingly used to indicate species' response to environmental stress across diverse taxa. Despite this broad use, few studies have explored telomere length in plants. Thus, evaluation of new approaches for measuring telomeres in plants is needed. Rapid advances in sequencing approaches and bioinformatic tools now allow estimation of telomere content from whole‐genome sequencing (WGS) data, a proxy for telomere length. While telomere content has been quantified extensively using quantitative polymerase chain reaction (qPCR) and WGS in humans, no study to date has compared the effectiveness of WGS in estimating telomere length in plants relative to qPCR approaches. In this study, we use 100Populusclones re‐sequenced using short‐read Illumina sequencing to quantify telomere length comparing three different bioinformatic approaches (Computel, K‐seek and TRIP) in addition to qPCR. Overall, telomere length estimates varied across different bioinformatic approaches, but were highly correlated across methods for individual genotypes. A positive correlation was observed between WGS estimates and qPCR, however, Computel estimates exhibited the greatest correlation. Computel incorporates genome coverage into telomere length calculations, suggesting that genome coverage is likely important to telomere length quantification when using WGS data. Overall, telomere estimates from WGS provided greater precision and accuracy of telomere length estimates relative to qPCR. The findings suggest WGS is a promising approach for assessing telomere length and, as the field of telomere ecology evolves, may provide added value to assaying response to biotic and abiotic environments for plants needed to accelerate plant breeding and conservation management.
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- Award ID(s):
- 1856450
- PAR ID:
- 10486111
- Publisher / Repository:
- Wiley-Blackwell
- Date Published:
- Journal Name:
- Molecular Ecology Resources
- Volume:
- 24
- Issue:
- 2
- ISSN:
- 1755-098X
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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