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Sanger sequencing

样品制备指南

DNA types	DNA length (including size)	template concentration,	total amount of template
Plasmid	< 6 kb	~50 ng/uL	~500 ng
	6 kb~10 kb	~80 ng/uL	~800 ng
	> 10 kb	~100 ng/uL	~1000 ng
Purified PCR products	<500 bp	~1 ng/uL
Unpurified PCR products	Original liquid total volume should be no less than 20uL
Bacterial culture	Direct extraction: 2-4 mL overnight cultured bacterial liquid if need to expand culture: >200 μL fresh bacterial liquid
Bacterial colonies on agar plates	When providing bacterial plates, ensure that the colonies are not too dense, and mark the colonies to be picked.
Sequencing primers	≥5 pmol/uL(5uM)	≥10 ul

Please provide the bacterial culture along with the type of antibiotics used. Commonly used antibiotics include ampicillin (Amp), kanamycin (Kan), chloramphenicol (Chl), etc. If non-standard antibiotics such as blasticidin are required, it is recommended to provide the corresponding antibiotics along with the sample.

For PCR products, if the fragment size is less than 100bp, sequencing after cloning is recommended. Please indicate the size of the bands and whether the bands are single.

The length of sequencing primers should be controlled within 20-30bp. Primers that are too short or too long can affect sequencing quality.

DNA samples provided should be dissolved in deionized water or double-distilled water, not in TE buffer.

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