How to use an Oxford Nanopore MinION to extract DNA from river water and determine which bacteria live in it PCR by 
Ciro Santilli 37 Updated 2025-07-16
Ciro Santilli 37 Updated 2025-07-16More generic PCR information at: Section "Polymerase chain reaction".
Because it is considered the less interesting step, and because it takes quite some time, this step was done by the event organizers between the two event days, so participants did not get to take many photos.
PCR protocols are very standard it seems, all that biologists need to know to reproduce is the time and temperature of each step.
This process used a Marshal Scientific MJ Research PTC-200 Thermal Cycler:
We added PCR primers for regions that surround the 16S DNA. The primers are just bought from a vendor, and we used well known regions are called 27F and 1492R. Here is a paper that analyzes other choices: academic.oup.com/femsle/article/221/2/299/630719 (archive) "Evaluation of primers and PCR conditions for the analysis of 16S rRNA genes from a natural environment" by Yuichi Hongoh, Hiroe Yuzawa, Moriya Ohkuma, Toshiaki Kudo (2003)
One cool thing about the PCR is that we can also add a known barcode at the end of each primer as shown at Code 1. "PCR diagram".
This means that we bought a few different versions of our 27F/1492R primers, each with a different small DNA tag attached directly to them in addition to the matching sequence.
This way, we were able to:
- use a different barcode for samples collected from different locations. This means we
- did PCR separately for each one of them
- for each PCR run, used a different set of primers, each with a different tag
- the primer is still able to attach, and then the tag just gets amplified with the rest of everything!
- sequence them all in one go
- then just from the sequencing output the barcode to determine where each sequence came from!
Input: Bacterial DNA (a little bit)
... --- 27S --- 16S --- 1492R --- ...
|||
|||
vvv
Output: PCR output (a lot of)
Barcode --- 27S --- 16S --- 1492RCode 1.
PCR diagram
. Finally, after purification, we used the Qiagen QIAquick PCR Purification Kit protocol to purify the generated from unwanted PCR byproducts.
Nick Leeson and the Fall of the House of Barings by Adam Curtis (1996) by Ciro Santilli 37 Updated 2025-07-16
Ciro Santilli 37 Updated 2025-07-16Tried a quick port to SQLite to get rid of annoying local databases for development, but failed, at c1c2cc4e448b279ff083272df1ac50d20c3304faandthen:fails with:Attempt to hack it:and after that it seems to run.
npm install sqlite3 --save-dev{
"type": "sqlite",
"database": "db.sqlite3",
"entities": ["src/**/**.entity{.ts,.js}"],
"synchronize": true
}npm startDataTypeNotSupportedError: Data type "timestamp" in "ArticleEntity.created" is not supported by "sqlite" database.--- a/src/article/article.entity.ts
+++ b/src/article/article.entity.ts
@@ -20,10 +20,10 @@ export class ArticleEntity {
@Column({default: ''})
body: string;
- @Column({ type: 'timestamp', default: () => "CURRENT_TIMESTAMP"})
+ @Column({ default: () => "CURRENT_TIMESTAMP"})
created: Date;
- @Column({ type: 'timestamp', default: () => "CURRENT_TIMESTAMP"})
+ @Column({ default: () => "CURRENT_TIMESTAMP"})
updated: Date;I can signup and login, terrible error reporting as usual, make sure to use long enough usernames/passwords.
However, article creation fails with:
Unhandled Rejection (TypeError): Cannot read property 'slug' of undefinedhello_world_len points to the special st_shndx == SHN_ABS == 0xF1FF.0xF1FF is chosen so as to not conflict with other sections.st_value == 0xD == 13 which is the value we have stored there on the assembly: the length of the string Hello World!.This is small optimization that our assembler does for us and which has ELF support.
The "last" gene, and also an E. Coli K-12 MG1655 gene of unknown function.
A collection of closely related and curated C. elegans datasets.
The default isolation level for SQLite is SERIALIZABLE
It does not appear possible to achieve the other two levels besides SERIALIZABLE and READ UNCOMMITED
Has Serial wire debug debug pre-soldered. Why would you ever get one without unless you are a clueless newbie like Ciro Santilli?!?!
It is however possible to solder it yourself on Raspberry Pi Pico W.
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