Martel, C. | Search bioRxiv

bioRxiv2022

Use of Solentim verified in-situ plate seeding (VIPSTM) enhances single-cell cloning efficiency

The primary goal in cell line development is to establish high-producer recombinant cell line(s) of single-cell origin. Traditionally, these cell lines are developed using limiting dilution cloning (LDC) method of single cell isolation, a rate-limiting, lengthy and labor-intensive process. The Verified-In-Situ-Plate-Seeding (VIPS) is an automated single-cell seeding and imaging equipment designed to accelerate cell line development workflow. In this study, VIPS was tested for efficiency and accuracy of single cell seeding in parallel with limiting dilution cloning (LDC). Three Chinese hamster ovary (CHO) derived cell lines with known clonal properties were tested under six different growth conditions (three growth media and two different kinds of microplates). Data showed VIPS and limiting dilution (LDC) have comparable cloning efficiency when CHO-M cells were tested. By contrast, the Verified In-Situ Plate Seeding (VIPS) produced 6-8-fold more clones of single-cell origin than LDC when CHO-K1 or CHO-S cells were tested. Moreover, the verified In-Situ Plate Seeding (VIPS) correctly identified single-cell and multiple-cells seeded wells with 65-72% and 52-81% accuracy, respectively. Taken together, the high throughput imaging and single-cell seeding capabilities of VIPS outperformed the rate-limiting LDC method and, therefore, has the potential to accelerate cell line development workflow.

cell biology↗

bioRxiv2021

Gender identity and sexual experience affect mating behaviour and chemical profile in the lesser mealworm, Alphitobius diaperinus (Coleoptera: Tenebrionidae)

Alphitobius diaperinus (Coleoptera: Tenebrionidae), the lesser mealworm, is one of the most significant pests of the poultry industry worldwide. These insects cause structural damage in poultry houses and transmit several diseases, impacting chickens productivity and rearing costs. Although semiochemicals may offer alternative insect pest management strategies, basic information regarding pheromone identity and their role on the behavioural ecology according to their circadian pattern of sexual behaviour of A. diaperinus is essentially lacking. This study is aimed to analyse the relation of gender identity and sexual experience of adults of A. diaperinus on their mating behaviour and whether this response is related to their CHC profiles secreted. The following steps were taken to achieve the studys goal. First, the circadian pattern of their sexual activity was observed in newly emerged pairs for at least twenty-one days (virgin adults) and experienced adults collected from the field to identify a difference based on their sexual experience and achieve the optimal mating season to develop the following assays. Subsequently, Y-tube olfactometer bioassays were conducted to evaluate their odour bouquet attraction based on gender and sexual experience. Additionally, mating behaviour bioassays were conducted to evaluate the two factor effects. Finally, cuticular analysis was performed using gas chromatography-mass spectrometry to evaluate possible chemical differences based on the two factors. With statistical and multivariate analysis, we found that behavioural, mating and chemical responses are different based on their sexual experience. The mating sequences were described into precopulatory, copulatory and postcopulatory phases. This finding gives us a deeper understanding of the sexual communication during mating. In summary, our findings provide new insights into the mating system and chemical ecology of A. diaperinus. The results presented here may serve as a base for further studies to develop strategies for managing this pest.

ecology↗