A new primer pair for barcoding of bees (Hymenoptera: Anthophila) without amplifying the orthologous coxA gene of Wolbachia bacteria

Objectives DNA barcoding became an effective method for the identification and monitoring of bees. However, standard primer pairs used for barcoding often result in (co-) amplification of bacterial endosymbionts of the genus Wolbachia, which are widespread among bee species. Here we designed a new primer pair and compared it with the performance of the standard Folmer-primers for a small sample set of bees representing the main taxonomic groups of bees. Results The newly designed primer pair (BeeCox1F1/BeeCox1R2) outperformed the standard barcoding primer (LCO1490/HCO2198). By generating barcodes for a small test set of bees we found that the new primer pair produced high-quality sequences in all cases for unambiguous species identification using BOLD. Conversely, the standard barcoding primers often co-amplified the homologous Wolbachia gene and resulted in mixed chromatogram signals. These sequences showed high similarity with the bacterial endosymbiont instead of the host.

Linking meta-omics to the kinetics of denitrification intermediates reveals pH-dependent causes of N2O emissions and nitrite accumulation in soil

Soil pH is a key controller of denitrification. We analysed the metagenomics/transcriptomics and phenomics of two soils from a long-term liming experiment, SoilN (pH 6.8) and un-limed SoilA (pH 3.8). SoilA had severely delayed N2O reduction despite early transcription of nosZ (mainly clade I), encoding N2O reductase, by diverse denitrifiers. This shows that post-transcriptionally hampered maturation of the NosZ apo-protein at low pH is a generic phenomenon. Identification of transcript reads of several accessory genes in the nos cluster indicated that enzymes for NosZ maturation were present across a range of organisms, eliminating their absence as an explanation for the failure to produce a functional enzyme. nir transcript abundances (for NO2− reductase) in SoilA suggest that low NO2− concentrations in acidic soils, often ascribed to abiotic degradation, are primarily due to biological activity. The accumulation of NO2− in neutral soil was ascribed to high nar expression (nitrate reductase). The -omics results revealed dominance of nirK over nirS in both soils while qPCR showed the opposite, demonstrating that standard primer pairs only capture a fraction of the nirK pool. qnor encoding NO reductase was strongly expressed in SoilA, implying an important role in controlling NO. Production of HONO, for which some studies claim higher, others lower, emissions from NO2− accumulating soil, was estimated to be ten times higher from SoilA than from SoilN. The study extends our understanding of denitrification-driven gas emissions and the diversity of bacteria involved and demonstrates that gene and transcript quantifications cannot always reliably predict community phenotypes.

A New Primer Pair for Barcoding Bees (Hymenoptera, Anthophila) Without Amplifying the Homologous Wolbachia Gene

ObjectivesDNA barcoding became an important method for the identification and monitoring of bees. However, standard primer pairs used for barcoding often result in (co-) amplification of bacterial endosymbionts of the genus Wolbachia, which are widespread among bee species. Here we designed a new primer pair and compared it with the performance of the standard Folmer-primers for small sample set of bees representing the main taxonomic groups of bees.ResultsThe newly designed primer pair (BeeCox1F1/BeeCox1R2) clearly outperformed the standard barcoding primer (LCO1490/HCO2198). By generating barcodes for a small test set of bees we found that the new primer pair produced in all cases clear sequences for unambiguous species identification using BOLD. In contrast, the standard barcoding primers often resulted in the amplification of the homologous Wolbachia gene, which either resulted in a mixed chromatogram signal or identification of the bacterial endosymbiont instead of the host.

Rancang Bangun Sistem Kalibrasi Alat Ukur Tekanan Rendah

Persaingan industri peralatan ukur sekarang sangatlah berat, hal ini<br />mengakibatkan mereka harus selalu mempertahankan atau meningkatkan kualitas produk<br />mereka. Untuk mempertahankan produk, mereka harus menggunakan peralatan-peralatan<br />ukur yang tepat dan ini adalah suatu keharusan. Seperti kita ketahui kalibrasi alat ukur<br />adalah salah satu cara untuk mempertahankan alat ukur tersebut berfungsi dengan baik.<br />Alat ukur tekanan seperti pressure gauge, pressure transmitter, dan alat-alat ukur tekanan<br />lain harus selalu dikalibrasi dan untuk alasan ini kita harus menggunakan standard primer<br />seperti Dead Weight Tester (DWT). Disebabkan karena ketelitian DWT yang sangat tinggi<br />maka pembuatan alat ukur ini sangatlah pelik karena harus ditunjang dengan permesinan<br />yang tinggi kepresisiannya. Pada penelitian ini kami telah membuat sebuah standard primer<br />tekanan dengan cara yang lebih sederhana, namun ketelitian dari instrumen tetap tinggi.<br />Instrumen ini dirancang dan didedikasikan untuk kalibrasi portable. Prinsip dasar dari alat<br />ukur ini adalah manometer pipa-U yang berisi cairan aquabides, dan merealisasikan<br />tekanan dengan persamaan P = ρ.g.Δh’. Densitas aquabides secara langsung dihitung<br />dengan menggunakan persamaan fungsi temperatur, sedangkan perbedaan tinggi dua<br />permukaan cairan diukur menggunakan Laser Displacement Sensor (LDS), dan nilai<br />percepatan gravitasi yang digunakan adalah 9.781369 m/s2. Pengujian alat ini dilakukan<br />dengan menghitung besarnya ketidakpastiaan instrumen dimana kita mendapatkan 0.18%<br />dari skala maksimum. Berdasarkan pengujian validasi yang dilakukan oleh Lembaga<br />Nasional Metrologi Indonesia (LIPI) alat ini bisa diterima sebagai standard primer.