title: Effects of sample age on data quality from targeted sequencing of museum specimens: what are we capturing in time?

authors: McGaughran, Angela

subjects: Science & Technology

abstract: Background: Next-generation sequencing (NGS) can recover DNA data from valuable extant and extinct museum specimens. However, archived or preserved DNA is difficult to sequence because of its fragmented, damaged nature, such that the most successful NGS methods for preserved specimens remain sub-optimal. Improving wet lab protocols and comprehensively determining the effects of sample age on NGS library quality are therefore of vital importance. Here, I examine the relationship between sample age and several indicators of library quality following targeted NGS sequencing of ~1300 loci using 271 samples of pinned moth specimens (Helicoverpa armigera) ranging in age from 5 to 117 years.

Results: I find that older samples have lower DNA concentrations following extraction and thus require a higher number of indexing PCR cycles during library preparation. When sequenced reads are aligned to a reference genome or to only the targeted region, older samples have a lower number of sequenced and mapped reads, lower mean coverage, and lower estimated library sizes, while the percentage of adapters in sequenced reads increases significantly as samples become older. Older samples also show the poorest capture success, with lower enrichment and a higher improved coverage anticipated from further sequencing.

Conclusions: Sample age has a significant, measurable impact on the quality of NGS data following targeted enrichment. However, incorporating an uracil-removing enzyme into the blunt-end repair step during library preparation could help to repair DNA damage, and using a method that prevents adapter-dimer formation may result in improved data yields.

text: Effects of sample age on data quality from targeted sequencing of museum specimens: what are we capturing in time? Background: Next-generation sequencing (NGS) can recover DNA data from valuable extant and extinct museum specimens. However, archived or preserved DNA is difficult to sequence because of its fragmented, damaged nature, such that the most successful NGS methods for preserved specimens remain sub-optimal. Improving wet lab protocols and comprehensively determining the effects of sample age on NGS library quality are therefore of vital importance. Here, I examine the relationship between sample age and several indicators of library quality following targeted NGS sequencing of ~1300 loci using 271 samples of pinned moth specimens (Helicoverpa armigera) ranging in age from 5 to 117 years.

Results: I find that older samples have lower DNA concentrations following extraction and thus require a higher number of indexing PCR cycles during library preparation. When sequenced reads are aligned to a reference genome or to only the targeted region, older samples have a lower number of sequenced and mapped reads, lower mean coverage, and lower estimated library sizes, while the percentage of adapters in sequenced reads increases significantly as samples become older. Older samples also show the poorest capture success, with lower enrichment and a higher improved coverage anticipated from further sequencing.

Conclusions: Sample age has a significant, measurable impact on the quality of NGS data following targeted enrichment. However, incorporating an uracil-removing enzyme into the blunt-end repair step during library preparation could help to repair DNA damage, and using a method that prevents adapter-dimer formation may result in improved data yields.

year: 2020-02-28

title: Reactions of isonitriles with [Fe₃(CO)₁₂] and [Ru₃(CO)₁₂] monitored by electrospray mass spectrometry: structural characterisation of [Fe₃(CO)₁₀(CNPh)₂] and [Ru₄(CO)₁₁(μ₃-η²-CNPh)₂(CNPh)]

authors: Decker, Corry

subjects: iron carbonyl

abstract: The reactions of [Fe₃(CO)₁₂] or [Ru₃(CO) ₁₂] with RNC (R=Ph, C₆H₄OMe-p or CH₂SO₂C₆H₄Me-p) have been investigated using electrospray mass spectrometry. Species arising from substitution of up to six ligands were detected for [Fe₃(CO)₁₂], but the higher-substituted compounds were too unstable to be isolated. The crystal structure of [Fe₃(CO)₁₀(CNPh)₂] was determined at 150 and 298 K to show that both isonitrile ligands were trans to each other on the same Fe atom. For [Ru₃(CO)₁₂] substitution of up to three COs was found, together with the formation of higher-nuclearity clusters. [Ru₄(CO)₁₁(CNPh)₃] was structurally characterised and has a spiked-triangular Ru₄ core with two of the CNPh ligands coordinated in an unusual μ₃-η² mode.
The substitution reactions of [M₃(CO)₁₂] by RNC have been investigated by electrospray mass spectrometry showing up to six COs can be replaced. [Fe₃(CO)₁₀(CNPh)₂] has both PhNC axially on the same Fe atom, and [Ru₄(CO)₁₁(μ₃-η²-CNPh)₂(CNPh)] has a spiked-triangular cluster core with two PhNC ligands in an unusual coordination mode.

text: Reactions of isonitriles with [Fe₃(CO)₁₂] and [Ru₃(CO)₁₂] monitored by electrospray mass spectrometry: structural characterisation of [Fe₃(CO)₁₀(CNPh)₂] and [Ru₄(CO)₁₁(μ₃-η²-CNPh)₂(CNPh)] The reactions of [Fe₃(CO)₁₂] or [Ru₃(CO) ₁₂] with RNC (R=Ph, C₆H₄OMe-p or CH₂SO₂C₆H₄Me-p) have been investigated using electrospray mass spectrometry. Species arising from substitution of up to six ligands were detected for [Fe₃(CO)₁₂], but the higher-substituted compounds were too unstable to be isolated. The crystal structure of [Fe₃(CO)₁₀(CNPh)₂] was determined at 150 and 298 K to show that both isonitrile ligands were trans to each other on the same Fe atom. For [Ru₃(CO)₁₂] substitution of up to three COs was found, together with the formation of higher-nuclearity clusters. [Ru₄(CO)₁₁(CNPh)₃] was structurally characterised and has a spiked-triangular Ru₄ core with two of the CNPh ligands coordinated in an unusual μ₃-η² mode.
The substitution reactions of [M₃(CO)₁₂] by RNC have been investigated by electrospray mass spectrometry showing up to six COs can be replaced. [Fe₃(CO)₁₀(CNPh)₂] has both PhNC axially on the same Fe atom, and [Ru₄(CO)₁₁(μ₃-η²-CNPh)₂(CNPh)] has a spiked-triangular cluster core with two PhNC ligands in an unusual coordination mode.

year: 2004-05-01

title: Representations of diversity in the revised New Zealand PictureBook Collection

authors: Daly, Nicola

subjects: picturebooks

abstract: Collections of children’s literature are used for a range of reasons including preservation, record keeping, and more recently the encouragement of intercultural understanding. Both the signifi- cance of learning about other cultures and the importance of children seeing themselves reflected in the books that they read have been discussed extensively. In 2010, the New Zealand PictureBook Collection (NZPBC) was developed in order to create a resource that reflected New Zealand national identity. In 2015 the NZPBC was revised and this article presents a visual and textual analysis of the diversity present in the sixty books nomi- nated through the lens of critical multicultural analysis, using four variables. Findings indicate considerable diversity is present in the collection in terms of representations of ethnicity and family; less so for representations of disability and languages.

text: Representations of diversity in the revised New Zealand PictureBook Collection Collections of children’s literature are used for a range of reasons including preservation, record keeping, and more recently the encouragement of intercultural understanding. Both the signifi- cance of learning about other cultures and the importance of children seeing themselves reflected in the books that they read have been discussed extensively. In 2010, the New Zealand PictureBook Collection (NZPBC) was developed in order to create a resource that reflected New Zealand national identity. In 2015 the NZPBC was revised and this article presents a visual and textual analysis of the diversity present in the sixty books nomi- nated through the lens of critical multicultural analysis, using four variables. Findings indicate considerable diversity is present in the collection in terms of representations of ethnicity and family; less so for representations of disability and languages.

year: 2017