Revisiting the impact of genomic hot spots: C12orf35 locus as a hot spot and engineering target.

Traditional Chinese hamster ovary (CHO) cell line development is based on random integration (RI) of transgene that causes clonal variation and subsequent large-scale clone screening. Therefore, site-specific integration (SSI) of transgenes into genomic hot spots has recently emerged as an alternative method for cell line development. However, the specific mechanisms underlying hot spot site formation remain unclear. In this study, we aimed to generate landing pad (LP) cell lines via the RI of transgenes encoding fluorescent reporter proteins flanked by recombination sites to facilitate recombinase-mediated cassette exchange. The RI-based LP cell line expressing high reporter levels with spontaneous C12orf35 locus deletion exhibited similar reporter fluorescent protein levels compared to targeted integrants with an identical reporter LP construct at the CHO genome hot spot, the C12orf35 locus. Additionally, Resf1, a C12orf35 locus gene, knockout (KO) in the RI-based LP cell line with conserved C12orf35 increased reporter expression levels, comparable to those in cell lines with C12orf35 locus disruption. These results indicate that the effect of SSI into the C12orf35 locus, a genomic hot spot, on high-level transgene expression was caused by C12orf35 disruption. In contrast to C12orf35 KO, KO at other well-known hot spot sites at specific loci of genes, including Fer1L4, Hprt1, Adgrl4, Clcc1, Dop1b, and Ddc, did not increase transgene expression. Overall, our findings suggest that C12orf35 is a promising engineering target and a hot spot for SSI-based cell line development.

Two new species of Cerviniella Smirnov, 1946 (Copepoda, Harpacticoida, Aegisthidae) from the Yellow Sea, Korea.

Two new species, Cerviniellabisegmentasp. nov. and C.permixtasp. nov., are described in detail with illustrations based on females from the Korean Yellow Sea. These species lacking the fourth leg endopod belong to the mirabilipes group, one of two species groups within the genus Cerviniella Smirnov, 1946. Both species can be distinguished from each other by the surface ornamentation of the cephalothorax, shape of the rostrum tip, antennule segments, armature formula of thoracic legs 1-4, and length ratio of the caudal rami. Cerviniellabisegmentasp. nov. is characterized by a short caudal ramus and a two-segmented antennary exopod, which are unique within the genus. Cerviniellapermixtasp. nov. differs from other congeners of the mirabilipes group by the seven-segmented antennule, the armature formulae of the exopod of the antenna and thoracic legs 1-4, and the modified apical inner element of the second endopodal segment of the second leg. The present study is the first to identify the genus Cerviniella in Korean waters, resulting in extension of its distribution area to East Asia.

Taxonomic review of the genus Stenocaris Sars (Copepoda, Harpacticoida, Cylindropsyllidae), with (re)descriptions of two Stenocaris species from the Far East.

The taxonomic concept of the genus Stenocaris Sars, 1909 is uncertain because none of the synapomorphies for the species of Stenocaris are defined. Detailed comparison of previous records of Stenocaris minor (T. Scott, 1892) from different localities reveals that this species represents a species complex composed of two species, S. minor s. str. and S. minor sensu Cottarelli & Venanzetti, 1989. Because the latter species has fundamental differences in the nature of the fifth leg in females and the sexual dimorphism of the second leg in males, we propose a new species for S. minor sensu Cottarelli & Venanzetti, 1989, S. figaroloensis sp. nov. We also suggest that S. minor sensu Apostolov, 1971, S. minor sensu Marinov, 1971, and S. minor sensu Apostolov & Marinov, 1988 from the Black Sea and S. minor sensu Wilson, 1932 from North America should be relegated to species inquirenda in the genus. Taxonomic review of the morphology of all Stenocaris species indicated that the generic concept must be restricted to accommodate S. minor s. str., S. gracilis Sars, 1909, S. intermedia Itô, 1972, S. figaroloensis sp. nov., and the South Korean new species, S. marcida sp. nov., based on the synapomorphic condition of the confluent fifth leg in males. As a result of our analysis, two Stenocaris species, S. baltica Arlt, 1983 and S. pygmaea Noodt, 1955, are transferred to the genus Vermicaris Kornev & Chertoprud, 2008 as V. baltica (Arlt, 1983) comb. nov. and V. pygmaea (Noodt, 1955) comb. nov. based on the synapomorphic characters of a reduced condition of the second and fifth legs. Additionally, S. arenicola Wilson, 1932 and S. kliei (Kunz, 1938) are allocated to a new genus, Huysicaris gen. nov., mainly characterized by obvious caudal rami with a recurved dorsal spinous process and convex inner margins, as H. arenicola (Wilson, 1932) comb. nov. and H. kliei (Kunz, 1938) comb. nov. A marine interstitial harpacticoid collected from the subtidal substrate off Dok-do Island in the East Sea of South Korea is proposed as S. marcida sp. nov. and the distribution of S. intermedia, originally known from its type locality in Japanese waters only, is extended to the East Sea of Korea and Russia. We provide their detailed descriptions and illustrations and discuss the morphological characters supporting their identities.

A new species of the genus Stylicletodes Lang, 1936 (Copepoda, Harpacticoida, Cletodidae) from South Korea.

Background: Marine benthic harpacticoid copepods are poorly known in Korea due to the difficulty in obtaining specimens. Currently, the genus Stylicletodes Lang, 1936, which is known to occur in subtidal sediments, has not been reported in this area so far. During surveys on the subtidal meiofauna, we found a new species of Stylicletodes from several subtidal muddy sediments in the Yellow Sea and South Sea. New information: In this study, we describe both sexes of a new species of Stylicletodes collected from the Yellow Sea and South Sea of Korea. Stylicletodestrifidus sp. nov. differs from its congeners in the following characteristics: the trifid rostrum, relative length ratio of the endopods to exopods on legs 1-4, reduced armature formulae on legs 3-4, constricted shape at mid-length of the anal somite, and structure of the sexually dimorphic male leg 3 with a two-segmented endopod. The new species underwent loss of the maxilliped, which is very rare in harpacticoids and is probably an important clue for the phylogeny of the species of Stylicletodes.

First report of the genus Bicorniphontodes (Copepoda, Harpacticoida, Ancorabolidae) in South Korea, with descriptions of three new species.

We report the occurrence of the genus Bicorniphontodes George, Glatzel & Schröder, 2019 in Korean waters, with descriptions of three new species: Bicorniphontodes lacuna sp. nov., B. comptus sp. nov., and B. huysi sp. nov. Morphology analysis was carried out to reconstruct the phylogenetic relationships of Bicorniphontodes species, including the three new species. Parsimony analysis based on 41 characters revealed that the three new species are clustered together as a monophyletic clade, of which B. horstgeorgei (George & Gheerardyn, 2015) is a sister species. The monophyletic status of three new species was supported by five synapomorphies, such as the micro-morphological conditions of the exopodal segments of the first leg, coxae of the second to fourth legs, exopod of the fifth leg in the female and baseoendopod of the fifth leg in the male, and the maxillular endopod represented by two setae. These three species can be easily distinguished based on the morphology of the rostrum, cephalothoracic processes, female genital double-somite, caudal rami, and second endopodal segment of the third leg in the male. The taxonomic position of B. bicornis sensu Kim, 2013 in Korean fauna was reevaluated based on the newly collected material from Udo Islet near Jeju Island; this resulted in a synonym of B. huysi sp. nov.

Taxonomy of three species of the genus Spinoncaea (Copepoda, Oncaeidae) in the North Pacific Ocean with focus on morphological variability.

Three species of Spinoncaea Böttger-Schnack, 2003 are newly recorded in three locations of the equatorial and temperate Pacific Ocean collected by using a net of 60 µm mesh size. For all three species, morphological characters and patterns of ornamentation were analyzed in detail and illustrations of both sexes, also including form variants of the females, are provided. For the first time, information about the variability of various continuous (morphometric) characters are given, such as the spine lengths on the rami of the swimming legs or the proportions of urosomites. The complementary morphological descriptions of the Pacific specimens focus on similarities or modifications of characters as compared to earlier descriptions of these species from the type locality and various other localities. For S. ivlevi (Shmeleva, 1966), originally but insufficiently described from the Adriatic Sea, the Pacific material is similar in most aspects to the comprehensive redescription of the species from the Red Sea and from the type locality, except for a difference in the morphometry of the distal endopod segment on the antenna, which is discussed here. For S. tenuis Böttger-Schnack, 2003, and S. humesi Böttger-Schnack, 2003, the Pacific material mostly coincides with the characteristic features as described in the original account from the Red Sea. For all three species, differences and/or additions in ornamentation details were found in Pacific specimens (e.g., on the intercoxal sclerite of the first swimming leg or on the genital somite of the male) and females with aberrant morphology were detected. Genetic analyses based on 12S srRNA revealed for two species, S. ivlevi and S. humesi, little or no differences in genetic sequences between Pacific specimens and those recorded from the Mediterranean Sea, thus demonstrating that specimens from both locations are conspecific. For S. tenuis, for which no comparable genetic data are available, 12S srRNA amplification was unsuccessful as was the amplification of mitochondrial COI (barcoding) for all three species. The applicability of using COI amplification for barcoding of oncaeid copepods is discussed.