Molecular biology

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Revision as of 13:14, 23 November 2010 (view source) Igchoi (Talk | contribs) (→=Gene replacement) ← Older edit		Latest revision as of 07:30, 16 January 2013 (view source) Csblsysop (Talk | contribs)
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-	{| align=left cellpadding=15	+	{| align=right cellpadding=15
	|__TOC__		|__TOC__
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	==Reagents & Protocols==		==Reagents & Protocols==
-	===recipes===	+	===Recipes===
-	*[[Chemicals]]	+	;*[[Chemicals]]
-	*[[media:mba.pdf|Commonly used reagents and equipment in molecular biology (PDF)]]	+	;*[[media:mba.pdf|Commonly used reagents and equipment in molecular biology (PDF)]]
-	*[[Concentration of antibiotics commonly used]]	+	;*[[Concentration of antibiotics commonly used]]
-	*For DNA extraction (to remove some protein contaminants & inactivate enzymes): Phenol:Chloroform=1:1(v:v) solution	+	;*For DNA extraction (to remove some protein contaminants & inactivate enzymes): Phenol:Chloroform=1:1(v:v) solution
-	*DNA sequencing check list	+	;*DNA sequencing check list
-	**[http://en.wikipedia.org/wiki/Phred_quality_score quality score]	+	:#[http://en.wikipedia.org/wiki/Phred_quality_score quality score]
-	**good sequencing result vs. bad sequencing result (see below example)	+	:#good sequencing result vs. bad sequencing result (see below example)
-	[[image:sequencingresult.jpg|left|thumb|good vs. bad sequencing example]]	+	::[[image:sequencingresult.jpg|center|thumb|good vs. bad sequencing example]]
	===Protocols===		===Protocols===
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	#Biological sequence editor [http://www.mbio.ncsu.edu/BioEdit/bioedit.html Bioedit]		#Biological sequence editor [http://www.mbio.ncsu.edu/BioEdit/bioedit.html Bioedit]
	#[http://en.wikipedia.org/wiki/Jmol Jmol]		#[http://en.wikipedia.org/wiki/Jmol Jmol]
		+	==Links==
		+	*[http://www.ncbe.reading.ac.uk/ncbe/protocols/ NCBE practical protocols]: very good illustration for undergraduate students

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Latest revision as of 07:30, 16 January 2013

Contents 1 Reagents & Protocols 1.1 Recipes 1.2 Protocols 1.2.1 LIC cloning 1.2.2 Conjugation 1.2.3 Gene replacement 2 Tools 2.1 webtools 2.2 softwares 3 Links

Reagents & Protocols

Recipes

• Chemicals
• Commonly used reagents and equipment in molecular biology (PDF)
• Concentration of antibiotics commonly used
• For DNA extraction (to remove some protein contaminants & inactivate enzymes): Phenol:Chloroform=1:1(v:v) solution
• DNA sequencing check list

1. quality score
2. good sequencing result vs. bad sequencing result (see below example)

good vs. bad sequencing example

Protocols

• PCR - General rules for primer design
• Genomic DNA extraction
• Competent cell preparation BSGC protocol (PDF)
  • Easy & rapid competent cell preparation reference link - by HJKo
• Electrocompetent cell Protocol (PDF)

LIC cloning

• Reference:

1. Error fetching PMID 2235490: [LIC]

• LIC cloning introduction LIC introduction - Ligation Independent Cloning
  • LIC cloning protocol (Berkeley Structural Genomics Center)
    • Vector preparation
    • Insert preparation
    • LIC transformation:
      • mix insert and vector in appropriate amounts
      • incubate 5~30 min(20 min) at room temperature
      • do transformation in E. coli (recommend to use electro-competent cells)

Conjugation

• Donor: E. coli S17-1

Gene replacement

• E. coli knockout by the Wanner method (Lambda Red Recombinase)

Tools

webtools

1. Oligo Tm calculation

softwares

1. NIH Image: Gel analysis software
2. Serial Cloner: DNA sequence management (PCR, restriction maps, etc.)
3. BioX (Mac OS X only)
4. Biological sequence editor Bioedit
5. Jmol

Links

• NCBE practical protocols: very good illustration for undergraduate students