Difference between revisions of "Part:BBa K2719009:Experience"

Revision as of 13:10, 17 October 2018

This experience page is provided so that any user may enter their experience using this part.
Please enter how you used this part and how it worked out.

Applications of BBa_K2719009

This part was cloned on pSB1C3 using EcoRI and PstI, later it was transformed on E.coli DH5a (Figure 1).

Then the plasmid was extracted and run on an agarose gel electrophoresis (Figure 2). Later an restriction was performed using NotI and was run on an agarose gel (Figure 3) to probe the presence of the insert. Later the extracted plasmid was transformed on E.coli BL21 (DE3).

The E.coli BL21 (DE3) colonies were later inoculated on liquid media (seed culture) and incubated overnight. From the seed culture 1 ml was inoculated on fresh LB+CAM broth, and at regular intervals the absorbance was measured. When the absorbance reached 0.7, IPTG was added to a final concentration of 1 mM and samples were taken after 5 and 16 h. Those samples were lysed. The lysate was loaded into a 18% polyacrylamide SDS-PAGE (Figure 4), from which a higher presence of protein was detected at 5 h after induction.

A western blot was made for this part and positive results were obtained. (Figure 5)

User Reviews

UNIQ6e513d0e7d01e78e-partinfo-00000000-QINU UNIQ6e513d0e7d01e78e-partinfo-00000001-QINU

Revision as of 13:10, 17 October 2018 (view source) AKARAM (Talk \| contribs) (→‎Applications of BBa_K2719009) ← Older edit		Revision as of 13:10, 17 October 2018 (view source) AKARAM (Talk \| contribs) (→‎Applications of BBa_K2719009) Newer edit →
Line 12:		Line 12:

	The <i>E.coli</i> BL21 (DE3) colonies were later inoculated on liquid media (seed culture) and incubated overnight. From the seed culture 1 ml was inoculated on fresh LB+CAM broth, and at regular intervals the absorbance was measured. When the absorbance reached 0.7, IPTG was added to a final concentration of 1 mM and samples were taken after 5 and 16 h. Those samples were lysed. The lysate was loaded into a 18% polyacrylamide SDS-PAGE (Figure 4), from which a higher presence of protein was detected at 5 h after induction.		The <i>E.coli</i> BL21 (DE3) colonies were later inoculated on liquid media (seed culture) and incubated overnight. From the seed culture 1 ml was inoculated on fresh LB+CAM broth, and at regular intervals the absorbance was measured. When the absorbance reached 0.7, IPTG was added to a final concentration of 1 mM and samples were taken after 5 and 16 h. Those samples were lysed. The lysate was loaded into a 18% polyacrylamide SDS-PAGE (Figure 4), from which a higher presence of protein was detected at 5 h after induction.
		+

	A western blot was made for this part and positive results were obtained. (Figure 5)		A western blot was made for this part and positive results were obtained. (Figure 5)