BBa_K2100050

BBa_K2100050 Version 1

Component

Source: http://parts.igem.org/Part:BBa_K2100050

Generated By: https://synbiohub.org/public/igem/igem2sbol/1

Created by: Trinh Nguyen

Date created: 2016-10-18 11:00:00

Date modified: 2016-10-23 12:49:37

pENTR L4_EGSH 2x k-turn_R1

Details

• Types: DnaRegion
• Roles: sequence_feature; DNA
• Sequences: BBa_K2100050_sequence (Version 1)

Description

This basic part entry vector is flanked by L4 and R1 sites, which are used to denote a promoter. This can be cascade with a gene (flanked by L1, L2 sites) using an LR reaction and cloned into a backbone that has a negative selection marker between R4 and R2 sites. This part adheres to RFC 65 for recombination based cloning of mammalian parts.

k-turn stands for kinh turn motif in the messenger RNA. The k-turn motif is a part of the L7Ae/k-turn RNA-based repressing system. By putting the k-turn motifs in pENTR promoter vector, the k-turns will be added into the 5'UTR of the gene after LR reaction. The k-turn adds in a higher gene regulation level to reduce the basal expression of the gene downstream due to leaky expression of the EGSH promoter

Notes

The distance from the k-turn repeats to the open reading frame of the downstream gene determines the efficiency of L7Ae/k-turn repressing system. Thus, we were considering adding k-turns right after the promoter (in the pENTR promoter vector) or right in front of the gene coding sequence (in the pENTR gene vector).

Source

EGSH was obtained using PCR extension from pENTR pEGSH (BBa_K2100021). The k-turn sequence was obtained from addgene (plasmids deposite by Dr. Ron Weiss' lab). We ordered the k-turns as single-stranded oligos with different Q sites for Golden Gate reaction.

Sequence Annotations

Sequence Annotation	Location	Component / Role(s)
pEGSH	1410,1931	promoter, feature/promoter
k-turn	1947,1969	non_covalent_binding_site, feature/binding
k-turn	1974,1996	non_covalent_binding_site, feature/binding
L4	1185,1281	sequence_feature, feature/dna
R1	2017,2145	feature/dna, sequence_feature

Other Properties

• igem#status: Planning
• synbiohub#ownedBy: user/james
• synbiohub#ownedBy: user/myers
• synbiohub#topLevel: BBa_K2100050/1

Member of these Collections

• iGEM Parts Registry