Team:UNIPV-Pavia/Material Methods/Measurements/Tecan/test16settembre

SEPTEMBER, 16TH

PLATE

EXPERIMENT DESCRIPTION

Motivation

The experiment was performed to evaluate the initiation transcription point (in term of absorbance) for self-inducible devices.

Methods

Inoculum into 1 ml LB + a suitable antibiotic of 8 ul of glycerol stock of:

• <partinfo>pSB1A2</partinfo> - <partinfo>BBa_K300024</partinfo> (wiki name: I7)
• <partinfo>pSB1A2</partinfo> - <partinfo>BBa_K300021</partinfo> (wiki name: I8)
• <partinfo>pSB1A2</partinfo> - <partinfo>BBa_K300022</partinfo> (wiki name: I9)
• <partinfo>pSB1A2</partinfo> - <partinfo>BBa_K300023</partinfo> (wiki name: I10)
• <partinfo>pSB1A2</partinfo> - <partinfo>BBa_K300019</partinfo> (wiki name: I12)
• <partinfo>pSB1A2</partinfo> - <partinfo>K173001</partinfo> (positive control)
• <partinfo>pSB1A2</partinfo> - <partinfo>BBa_B0032</partinfo> (negative control)
• <partinfo>pSB4C5</partinfo> - <partinfo>BBa_K300030</partinfo> (wiki name I14) co-transformed with <partinfo>pSB1A3</partinfo> - <partinfo>BBa_T9002</partinfo>
• <partinfo>pSB4C5</partinfo> - <partinfo>BBa_K300028</partinfo> (wiki name I15) co-transformed with <partinfo>pSB1A3</partinfo> - <partinfo>BBa_T9002</partinfo>
• <partinfo>pSB4C5</partinfo> - <partinfo>BBa_K300029</partinfo> (wiki name I16) co-transformed with <partinfo>pSB1A3</partinfo> - <partinfo>BBa_T9002</partinfo>
• <partinfo>pSB4C5</partinfo> - <partinfo>BBa_K300025</partinfo> (wiki name I17) co-transformed with <partinfo>pSB1A3</partinfo> - <partinfo>BBa_T9002</partinfo>
• <partinfo>pSB4C5</partinfo> - <partinfo>BBa_K300026</partinfo> (wiki name I18) co-transformed with <partinfo>pSB1A3</partinfo> - <partinfo>BBa_T9002</partinfo>
• <partinfo>pSB4C5</partinfo> - <partinfo>BBa_K300027</partinfo> (wiki name I19) co-transformed with <partinfo>pSB1A3</partinfo> - <partinfo>BBa_T9002</partinfo>
• <partinfo>pSB1A3</partinfo> - <partinfo>BBa_T9002</partinfo> (negative control)
• <partinfo>pSB4C5</partinfo> - <partinfo>BBa_K300024</partinfo> (wiki name: I7-4C5)
• <partinfo>pSB4C5</partinfo> - <partinfo>BBa_K300021</partinfo> (wiki name: I8-4C5)
• <partinfo>pSB4C5</partinfo> - <partinfo>BBa_K300023</partinfo> (wiki name: I10-4C5)
• <partinfo>pSB4C5</partinfo> - <partinfo>BBa_K300019</partinfo> (wiki name: I12-4C5)

They were let grow ON at +37°C, 220 rpm.

The following day cultures were diluted 1:100 in a final volume of 1 ml and let grow again for about five hours at +37°C, 220 rpm. Then cultures were centrifuged at 2000 rpm, 25°C, the supernatants were removed and the pellets were resuspended. Optical density (O.D.) of each cell culture was than measured with TECAN Infinte F200. Samples were diluted in order to obtain the same O.D. equal to 0,02. Then O.D. and green Fluorecence were measured.

RESULTS

Cultures resulted immediately induced because of the high concentration of homoserine-lactone that remained in the solutions. This prevented from calculating the ODstart.

Tecan Test