Customize cgh-1 Antibody

Description

Antibody Generation

cgh-1 antibodies are typically polyclonal, raised against specific peptides of the CGH-1 protein. Key production methods include:

Peptide Selection:
- Rabbit antisera were generated against the peptide CDPKLYVADQQLVDAADETTA (residues 411–431) .
- Chicken antibodies were raised against a CAR-1 peptide (246–265), though these are less relevant to CGH-1 studies .
Purification:
- Affinity purification using Sulfolink resin or protein L/G agarose .
- Validation via western blotting and immunoprecipitation (IP) to confirm specificity .

Key Validation Data

Method	Purpose	Outcome
Western Blotting	Confirm germline-specific expression	Detects a 45 kDa band in wild-type lysates, absent in glp-4(bn2) mutants .
Immunoprecipitation	Identify CGH-1 interaction partners	Co-purifies CAR-1, CEY-2, CEY-3, CEY-4, and DCAP-2 .
Immunofluorescence	Localize CGH-1 in germ cells/embryos	Detects CGH-1 in P-granules and cytoplasmic foci .

Immunoprecipitation (IP)

cgh-1 antibodies are essential for isolating CGH-1-containing ribonucleoprotein (RNP) complexes:

RNA-Dependent Interactions:
- Co-IP with CAR-1, CEY-2, and CGH-1 requires RNA integrity . RNase treatment disrupts these interactions .
Functional Partners:
- CGH-1 associates with DCAP-2 (decapping enzyme) in somatic P bodies and ATX-2/PAB-1 (translational regulators) in germ cells .

Immunofluorescence

Used to map CGH-1 localization:

Germline: Concentrates in P-granules (germ cell precursors) and cytoplasmic particles in the gonad rachis .
Embryos: Localizes to interzonal microtubules during anaphase, critical for cytokinesis .

RNA Immunoprecipitation (RIP-Chip)

Identifies CGH-1-bound mRNAs:

Germ Cell Storage Bodies: CGH-1 binds and stabilizes maternal mRNAs (e.g., pos-1) in oocytes, protecting them from degradation .
Somatic P Bodies: Associates with decapping machinery (DCAP-1/2, PATR-1) in somatic cells .

Role in Cytokinesis

Anaphase Defects: Depletion of CGH-1 via cgh-1(ok492) mutants disrupts interzonal microtubule bundles, causing cytokinesis failure in embryos .
CAR-1 Dependency: CGH-1 controls CAR-1 localization; cgh-1(RNAi) worms show mislocalized CAR-1 and defective spindle assembly .

Regulation of Maternal mRNAs

Stabilization: CGH-1 storage bodies protect ~100 maternal mRNAs from degradation during oogenesis, as shown by RIP-Chip and cgh-1 mutant analysis .
Differential Functions:

Tissue CGH-1 Complex Function
Somatic P bodies (with PATR-1) mRNA decapping and degradation .
Germ Cells Storage bodies mRNA storage and protection .

Tissue	CGH-1 Complex	Function
Somatic	P bodies (with PATR-1)	mRNA decapping and degradation .
Germ Cells	Storage bodies	mRNA storage and protection .

Apoptosis and Germ Cell Homeostasis

Physiological Apoptosis: CGH-1 suppresses apoptosis in oocytes; cgh-1(RNAi) causes ~100% oocyte loss .
Compensatory Mechanisms: Increased apoptosis in cgh-1 mutants may mitigate oogenesis defects by eliminating compromised germ cells .

Product Specs

Buffer

Preservative: 0.03% ProClin 300; Constituents: 50% Glycerol, 0.01M PBS, pH 7.4

Form

Liquid

Lead Time

14-16 weeks (made-to-order)

Synonyms

cgh-1 antibody; C07H6.5ATP-dependent RNA helicase cgh-1 antibody; EC 3.6.4.13 antibody; Conserved germline helicase 1 antibody

Target Names

cgh-1

Uniprot No.

Q95YF3

Target Background

Function

CGH-1 is a probable RNA helicase essential for oocyte and sperm function. Its crucial role lies in preventing the physiological germline apoptosis that would otherwise eliminate nearly all developing oocytes.

Gene References Into Functions

In developing oocytes, CGH-1 particles accumulate specifically in response to meiotic development. PMID: 15986473
The regulation of physiological germ cell apoptosis is significantly influenced by specific functions of the CGH-1/CAR-1 RNA-protein complex. PMID: 16221731
Studies have identified distinct somatic and germ cell CGH-1 functions, differentiated by the involvement of PATR-1. Furthermore, during oogenesis, numerous translationally regulated mRNAs are specifically protected by a CGH-1-dependent mechanism. PMID: 18695045

Database Links

KEGG: cel:CELE_C07H6.5

STRING: 6239.C07H6.5.2

UniGene: Cel.17052

Protein Families

DEAD box helicase family, DDX6/DHH1 subfamily

Subcellular Location

Cytoplasm.

Tissue Specificity

Expression is restricted to two germline precursors Z2 and Z3 in L1 stage hermaphrodites, and is detectable specifically in the gonad at low levels into the L3 stage. Expression is significantly higher during the early L4 stage. In adults, expression rema

Q&A

FAQs for Researchers Investigating CGH-1 Antibody in Academic Research

Advanced Research Questions

How to resolve contradictions in CGH-1’s role in mRNA degradation vs. protection?
- Context-dependent analysis:
  - In somatic cells, CGH-1 promotes decapping via PATR-1 and DCAP-2 .
  - In oocytes, CGH-1 stabilizes maternal mRNAs by sequestering them from decapping machinery .
- Strategy: Compare transcript stability (e.g., RNA-seq) in cgh-1(RNAi) vs. patr-1(RNAi) mutants.
What experimental designs optimize co-staining with CGH-1 and other P-body markers?
- Combination markers:
  
  Marker Compatibility with CGH-1 Functional Overlap
  CAR-1 High (RNase-sensitive) mRNA storage
  DCAP-2 Moderate (somatic-specific) Decapping
  WAGO-1 Low (germline-specific) Argonaute pathways
- Recommendation: Use sequential staining with antibody cross-absorption to minimize overlap.
How to assess CGH-1’s mechanistic role in germline apoptosis?
- Approach:
  - Quantify apoptosis in cgh-1(RNAi) mutants using acridine orange staining .
  - Combine with translational reporters (e.g., gld-1::GFP) to link mRNA regulation to cell survival.
- Key data: cgh-1 depletion increases physiological germline apoptosis from ~50% to >90% .
What are the limitations of CGH-1 antibody in detecting post-translational modifications?
- Challenge: No commercial antibodies target phosphorylated or ubiquitinated CGH-1.
- Workaround: Pair IP with mass spectrometry to identify modifications in immunoprecipitated complexes .

Marker	Compatibility with CGH-1	Functional Overlap
CAR-1	High (RNase-sensitive)	mRNA storage
DCAP-2	Moderate (somatic-specific)	Decapping
WAGO-1	Low (germline-specific)	Argonaute pathways

Methodological Troubleshooting

How to address cross-reactivity with paralogs like GLH-1/GLH-4?
- Solution:
  - Use glh-1;glh-4 double mutants as negative controls .
  - Validate antibody specificity via CRISPR-Cas9 knockout lines (e.g., cgh-1(tmXXXX)) .
How to quantify CGH-1 dynamics under stress conditions (e.g., oxidative stress)?
- Protocol:
  1. Treat with paraquat (10 mM, 1 hr) to induce stress.
  2. Image CGH-1 granules via super-resolution microscopy.
  3. Use particle analysis software (e.g., ImageJ) to measure granule size/number.
- Expected outcome: Stress increases somatic P-body size by 2–3× .

Data Interpretation Frameworks

Integrating CGH-1 interaction data with transcriptomic datasets:
- Workflow:
  1. Perform RIP-seq (RNA immunoprecipitation sequencing) with CGH-1 antibody.
  2. Overlap with CLIP-seq data from partners like CAR-1 or WAGO-1 .
  3. Use pathway analysis (e.g., DAVID) to identify enriched biological processes.
- Example finding: CGH-1 binds mRNAs encoding translational regulators (e.g., pab-1, atx-2) .

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cgh-1 Antibody