Germ line transformation and in vivo labeling of nuclei in Diptera: report on Megaselia abdita (Phoridae) and Chironomus riparius (Chironomidae)

Vitamin B12 deficiency is common in people of all ages who consume a low intake of animal-source foods, including populations in developing countries. It is also prevalent among the elderly, even in wealthier countries, due to their malabsorption of B12 from food. Several methods have been applied to diagnose vitamin B12 malabsorption, including Schillings test, which is now used rarely, but these do not quantify percent bioavailability. Most of the information on B12 bioavailability from foods was collected 40 to 50 years ago, using radioactive isotopes of cobalt to label the corrinoid ring. The data are sparse, and the level of radioactivity required for in vivo labeling of animal tissues can be prohibitive. A newer method under development uses a low dose of radioactivity as 14C-labeled B12, with measurement of the isotope excreted in urine and feces by accelerator mass spectrometry. This test has revealed that the unabsorbed vitamin is degraded in the intestine. The percent bioavailability is inversely proportional to the dose consumed due to saturation of the active absorption process, even within the range of usual intake from foods. This has important implications for the assessment and interpretation of bioavailability values, setting dietary requirements, and interpreting relationships between intake and status of the vitamin.

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Germ-line transformation and RNAi of the ladybird beetle, Harmonia axyridis

Insect Molecular Biology ◽

10.1111/j.1365-2583.2006.00665.x ◽

2006 ◽

Vol 15 (4) ◽

pp. 507-512 ◽

Cited By ~ 26

Author(s):

H. Kuwayama ◽

T. Yaginuma ◽

O. Yamashita ◽

T. Niimi

Keyword(s):

Harmonia Axyridis ◽

Germ Line ◽

Ladybird Beetle ◽

Germ Line Transformation

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Monitoring of changes in the membrane proteome during stationary phase adaptation of Bacillus subtilis using in vivo labeling techniques

PROTEOMICS ◽

10.1002/pmic.200701081 ◽

2008 ◽

Vol 8 (10) ◽

pp. 2062-2076 ◽

Cited By ~ 48

Author(s):

Annette Dreisbach ◽

Andreas Otto ◽

Dörte Becher ◽

Elke Hammer ◽

Alexander Teumer ◽

...

Keyword(s):

Bacillus Subtilis ◽

Stationary Phase ◽

Membrane Proteome ◽

In Vivo Labeling

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Metabolic in Vivo Labeling Highlights Differences of Metabolically Active Microbes from the Mucosal Gastrointestinal Microbiome between High-Fat and Normal Chow Diet

Journal of Proteome Research ◽

10.1021/acs.jproteome.6b00973 ◽

2017 ◽

Vol 16 (4) ◽

pp. 1593-1604 ◽

Cited By ~ 19

Author(s):

Andreas Oberbach ◽

Sven-Bastiaan Haange ◽

Nadine Schlichting ◽

Marco Heinrich ◽

Stefanie Lehmann ◽

...

Keyword(s):

Chow Diet ◽

High Fat ◽

Gastrointestinal Microbiome ◽

In Vivo Labeling ◽

Normal Chow ◽

Normal Chow Diet

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Probing Drosophila gene function by antisense RNA

Genome ◽

10.1139/g89-063 ◽

1989 ◽

Vol 31 (1) ◽

pp. 422-425 ◽

Cited By ~ 5

Author(s):

Reinhard Schuh ◽

Herbert Jäckle

Keyword(s):

Antisense Rna ◽

Germ Line ◽

Conventional Technique ◽

Transcription Unit ◽

The Body ◽

Mutant Phenotype ◽

Drosophila Embryo ◽

Alternative Approach ◽

Pattern Forming ◽

Germ Line Transformation

The conventional technique for assigning a particular genetic function to a cloned transcription unit has relied on the rescue of the mutant phenotype by germ line transformation. An alternative approach is to mimic a mutant phenotype by the use of antisense RNA injections to produce phenocopies. This approach has been successfully used to identify genes involved in early pattern forming processes in the Drosophila embryo. At the time when antisense RNA is injected, the embryo develops as a syncytium composed of about 5000 nuclei which share a common cytoplasm. The gene interactions required to establish the body plan occur before cellularization at the blastoderm stage. Thus the nuclei and their exported transcripts are accessible to the injected antisense RNA. The antisense RNA interferes with the endogenous RNA by an as yet unidentified mechanism. The extent of interference is only partial and produces phenocopies with characteristics of weak mutant alleles. In our lab and others, this approach has been successfully used to identify several genes required for normal Drosophila pattern formation.Key words: Drosophila segmentation, phenocopy, antisense RNA, Krüppel gene.

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Delimiting regulatory sequences of the Drosophila melanogaster Ddc gene

Molecular and Cellular Biology ◽

10.1128/mcb.6.12.4548-4557.1986 ◽

1986 ◽

Vol 6 (12) ◽

pp. 4548-4557

Author(s):

J Hirsh ◽

B A Morgan ◽

S B Scholnick

Keyword(s):

Drosophila Melanogaster ◽

Germ Line ◽

Regulatory Elements ◽

P Element ◽

Developmental Expression ◽

Upstream Region ◽

Regulatory Sequences ◽

Flanking Sequences

We delimited sequences necessary for in vivo expression of the Drosophila melanogaster dopa decarboxylase gene Ddc. The expression of in vitro-altered genes was assayed following germ line integration via P-element vectors. Sequences between -209 and -24 were necessary for normally regulated expression, although genes lacking these sequences could be expressed at 10 to 50% of wild-type levels at specific developmental times. These genes showed components of normal developmental expression, which suggests that they retain some regulatory elements. All Ddc genes lacking the normal immediate 5'-flanking sequences were grossly deficient in larval central nervous system expression. Thus, this upstream region must contain at least one element necessary for this expression. A mutated Ddc gene without a normal TATA boxlike sequence used the normal RNA start points, indicating that this sequences is not required for start point specificity.

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Fetal liver pro-B and pre-B lymphocyte clones: expression of lymphoid-specific genes, surface markers, growth requirements, colonization of the bone marrow, and generation of B lymphocytes in vivo and in vitro

Molecular and Cellular Biology ◽

10.1128/mcb.12.2.518-530.1992 ◽

1992 ◽

Vol 12 (2) ◽

pp. 518-530

Author(s):

R Palacios ◽

J Samaridis

Keyword(s):

B Cell ◽

B Lymphocytes ◽

Stromal Cells ◽

B Lymphocyte ◽

Germ Line ◽

Fetal Liver ◽

Rag 1 ◽

B Cell Precursors

We describe here the development and characterization of the FLS4.1 stromal line derived from 15-day fetal liver of BALB/c embryos and defined culture conditions that efficiently support the cloning and long-term growth of nontransformed B-220+ 14-day fetal liver cells at two stages of B-cell development, namely, pro-B lymphocytes (immunoglobulin [Ig] genes in germ line configuration) and pre-B cells (JH-rearranged genes with both light-chain Ig genes in the germ line state). All B-cell precursor clones require recombinant interleukin-7 (rIL-7) and FLS4.1 stromal cells for continuous growth in culture, but pro-B lymphocyte clones can also proliferate in rIL-3. None proliferate in rIL-1, rIL-2, rIL-4, rIL-5, rIL-6, or leukemia inhibitory factor. FLS4.1 stromal cells synthesize mRNA for Steel factor but not for IL-1 to IL-7; all pro-B and pre-B clones express c-Kit, the receptor for Steel factor, and a c-Kit-specific antibody inhibits the enhanced proliferative response of fetal liver B-220+ B-cell precursors supported by FLS4.1 stromal cells and exogenous rIL-7 but does not affect that promoted by rIL-7 alone. Northern (RNA) blot analysis of the expression of the MB-1, lambda 5, Vpre-B, c mu, RAG-1, and RAG-2 genes in pro-B and pre-B clones show that transcription of the MB-1 gene precedes IgH gene rearrangement and RNA synthesis from c mu, RAG-1, RAG-2, lambda 5, and Vpre-B genes. All clones at the pre-B-cell stage synthesize mRNA for c mu, RAG-1, and RAG-2 genes; transcription of the lambda 5 and Vpre-B genes seems to start after D-to-JH rearrangement in B-cell precursors, indicating that the proteins encoded by either gene are not required for B-cell progenitors to undergo D-to-JH gene rearrangement. These findings mark transcription of the MB-1 gene as one of the earliest molecular events in commitment to develop along the B-lymphocyte pathway. Indeed, both pro-B and pre-B clones can generate in vitro and in vivo B lymphocytes but not T lymphocytes; moreover, these clones do not express the CD3-gamma T-cell-specific gene, nor do they have rearranged gamma, delta, or beta T-cell antigen receptor genes.

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