🎯 Objective
To determine whether the genome of S. epidermidis HD46 contains known genes associated with natural cadmium resistance.
✅ Step 1: Prepare the Genome File
- Confirm that the genome file is in a compatible format such as
.fasta, .gbk, or .gff + .fna.
- If the genome is not annotated, proceed to annotation.
✅ Step 2: Annotate the Genome (if needed)
Option 1: Prokka (command-line tool)
Use Prokka to generate gene predictions and protein sequences.
Example command:
bash
prokka HD46.fasta --outdir prokka_out --prefix HD46
Option 2: RAST (web-based tool)
If you prefer a graphical interface:
- Go to RAST Server
- Create a free account and log in
- Upload your genome in FASTA format
- Select the default annotation pipeline
- Wait for the annotation to complete (this may take some time)
- Download the annotated protein sequences (.faa) and genome features (.gff, .tbl, etc.)
✅ Step 3: Identify Known Cadmium Resistance Genes
Look for genes known to confer cadmium resistance, including:
- cadA: a cadmium-translocating P-type ATPase
- cadC: regulatory protein for cad operon
- czcA / czcD: part of heavy metal efflux systems
- arsRBC operon: primarily arsenic-related but may overlap in function
- copA: copper ATPase, sometimes associated with cadmium resistance
✅ Step 4: Search for Homologous Genes Using BLAST
Create a local BLAST database
Convert the annotated protein sequences into a searchable format:
bash
makeblastdb -in HD46.faa -dbtype prot
Prepare a FASTA File with Reference Cadmium Resistance Proteins
- Download cadA, cadC, and other related protein sequences from NCBI or UniProt.
- Save them into a single FASTA file named
cadmium_genes.faa.
Run BLASTP:
bash
blastp -query cadmium_genes.faa -db HD46.faa -evalue 1e-5 -outfmt 6 -out cadmium_blast_results.txt
- Review the output for significant matches based on:
- Percent identity
- E-value
- Alignment length
✅ Step 5: Use the BacMet Database
BacMet is a curated database of experimentally verified metal resistance genes.
- Go to the BacMet download page
- Download the BacMet2 protein FASTA file.
Create a local BLAST database:
bash
makeblastdb -in BacMet2_PROTEIN.fasta -dbtype prot
Run BLASTP:
bash
blastp -query HD46.faa -db BacMet2_PROTEIN.fasta -evalue 1e-5 -outfmt 6 -out bacmet_hits.txt
- Analyze the output to identify any matches to known cadmium resistance genes.
✅ Step 6: Optional - Analyze with CARD
Although CARD primarily focuses on antibiotic resistance, some metal resistance genes are included.
- Visit the CARD RGI tool
- Upload your
.faa file (e.g., HD46.faa)
- Review the hits for any relevant metal resistance gene annotations
✅ Step 7: Visual Inspection of Gene Clusters
Use a genome browser like Artemis, Geneious, or SnapGene to open the annotated genome.
Look for:
- Clusters of metal resistance genes (e.g., cadA, cadC, czcA/D)
- Regulatory genes upstream (e.g., cadC before cadA)
- Adjacent stress response or membrane transport genes
These patterns may suggest operon-like organization and co-regulation.
✅ Step 8: Domain Confirmation
To verify if candidate genes are functionally related to cadmium resistance:
Look for domains such as:
- Heavy-metal-associated (HMA) domains
- P-type ATPase domains
- ArsR-type helix-turn-helix regulators
✅ Step 9: Interpret Results
- A strong match to cadA or cadC with conserved domains indicates cadmium resistance potential.
- Presence of gene clusters supports likely functionality.
- Distant or partial homologs may require experimental validation.
- Absence of canonical genes doesn't exclude alternative mechanisms of resistance.
🔧 Tools Mentioned
- Prokka – Command-line annotation
- RAST – Web-based annotation
- BLAST+ – Protein similarity search
- BacMet – Metal resistance gene reference
- CARD – Antibiotic and metal resistance database
- InterProScan / CD-Search – Domain analysis
- Artemis / Geneious – Genome browsing and visualization
镉抗性基因
镉抗性基因是指能够使生物体（如细菌、植物或其他微生物）在含有有毒重金属镉的环境中生存和繁殖的基因。这些基因通常编码一些蛋白质，帮助细胞排除、解毒或固定镉离子，从而减轻镉的毒性影响。
镉抗性基因的主要机制：
外排泵（Efflux pumps）
这类蛋白能够将镉离子主动泵出细胞外，降低细胞内镉的浓度。例如，cadA基因编码一种ATP驱动的镉离子外排蛋白。
金属结合蛋白
如金属硫蛋白（metallothionein），可以结合镉离子，使其失去毒性。
调控蛋白
这些蛋白调节镉抗性基因的表达，确保在有镉存在时激活抵抗机制。
典型例子：
cadA基因
最早在金黄色葡萄球菌的质粒中发现，能编码一种专门将镉离子泵出细胞的蛋白。
czc系统
存在于某些细菌中（如金属耐受铜绿假单胞菌），可以同时抵抗镉、锌和钴的外排系统。
金属硫蛋白
广泛存在于植物和微生物中，能结合镉离子，防止其对细胞的损伤。
如果你想了解更详细的基因序列、具体的生物种类或者镉抗性的应用，告诉我，我可以帮你进一步说明！
Cadmium 意思
“Cadmium” 的意思是 镉，是一种化学元素，符号是 Cd，原子序数是 48。
- 性质：银白色的金属，有延展性和柔软性。
- 毒性：镉是一种有毒的重金属，对人体和环境都有害，长期接触会导致中毒。
- 用途：主要用于制造镉镍电池、电镀和颜料等。
所以，“Cadmium” 就是指这种元素“镉”。