vnti11 basics course

basics course

Introduction

Introduction

Database Explorer

Molecule Display

Molecule Editing

Restriction Enzymes

Primer Design and PCR analysis

Molecule construction (cloning)

Introduction

Download VectorNTI11 from Invitrogen

http://www.invitrogen.com/site/us/en/home/LINNEA-Online-Guides/

LINNEA-Communities/Vector-NTI-Community/Vector-NTI.html

After installation the software will run in DEMO mode

until it is unlocked by a static or dynamic license.

Dynamic licenses are available by VIB (BITS).

Overview modules

VectorNTI Explorer:

The database to store your data

VectorNTI:

Sequence creation, mapping, analysis,

design, annotation, illustration and

molecular biology data management

VectorNTI Explorer

VectorNTI Explorer

New Object

VectorNTI Explorer

Organize your objects into groups using subsets

(and separators)

New Subset

VectorNTI Explorer

Organize your objects into groups using subsets

separator

Subset

VectorNTI Explorer

New Object

VectorNTI Explorer

VectorNTI Explorer

All molecules from all subsets are visible under MAIN

NEWMOL is in the MAIN set, not under any subset

VectorNTI Explorer

All molecules from all subsets are visible under MAIN

NEWMOL is in the MAIN set, not under any subset

Place NEWMOL in a subset by clicking, holding and moving the

name under MAIN to the subset name (DRAG and DROP)

VectorNTI Explorer

Subsets and separators can be moved in a similar way

DELETE = delete from database

DISMISS = remove subset (molecules NOT removed)

CLEAR – EXCLUDE = remove molecules from subset, NOT from MAIN database

VectorNTI Explorer


DELETE = delete from database (= pressing ‘delete’!)



VectorNTI Explorer


DELETE = delete from database



VectorNTI Explorer

subset selected

molecule selected

VectorNTI Explorer

VectorNTI Explorer

We want to gather all NCBI molecules in a separate subset for easy access

VectorNTI Explorer

VectorNTI Explorer

Backup on other location

e.g. network drive or external drive

Local database on network drive

» use same database on ≠ computers

We want to add some more structured information to each entry -> new field!

VectorNTI Explorer

New field Import fields

VectorNTI Explorer

VectorNTI ExplorerPaste the data

in e.g. Excel

VectorNTI Explorer

Summary

» Create, edit and delete objects and subsets

» Organize objects into convenient groups (subsets)

» Import and export objects and archives

» Perform database searches

VectorNTI Explorer

Exercises

» Import restriction enzymes from REBASE

http://rebase.neb.com (bairoch.nnn file)

» Add a new DNA marker in the Gel Markers database

» Select the pUC plasmids and export them into an archive file

» Make 2 RE subsets:

(1) BamHI, EcoRI, EcoRV, HindIII, NcoI,

NdeI, PstI, PvuI, SmaI and XmaI

(2) 6bp non-ambiguous

VectorNTI Explorerhttp://Rebase.neb.com/rebase/rebase.html

VectorNTI Explorer

VectorNTI Explorer

Save file as...

» bairoch.nnnR

VectorNTI Explorer

Exercises

» Import restriction enzymes from REBASE

http://rebase.neb.com (bairoch.nnn file)

» Add a new DNA marker in the Gel Markers database

» Select the pUC plasmids and export them into an archive file

» Make 2 RE subsets:

(1) BamHI, EcoRI, EcoRV, HindIII, NcoI,

NdeI, PstI, PvuI, SmaI and XmaI

(2) 6bp non-ambiguous

VectorNTI

Start VectorNTI...

VectorNTI

...

MAIN MENU

Links to the other modules of the Vector NTI suite

VectorNTI

...

MAIN TOOLBARChangeable toolbar

Depending on

which pane is

active

VectorNTI

...TEXT WINDOW

VectorNTI

... GRAPHICS WINDOW

VectorNTI

...

SEQUENCE WINDOW

VectorNTI

...

Local

Database

VectorNTI

...

Open the local

database

VectorNTI

...

Click pACYC177 to open

VectorNTI

...

Resize windows

for a better view

VectorNTI

...

VectorNTI

...

Copy complete

graphics

1

VectorNTI

...

VectorNTI

...

but COPY (Ctrl+C)

& PASTE (Ctrl+V)

also works...

VectorNTI

...

VectorNTI

...

Open the local

database

VectorNTI

... Click pBR322 to open

VectorNTI

...

VectorNTI

...

Molecule display setup

VectorNTI

...

VectorNTI

...

How to edit your

molecule

VectorNTI

...

VectorNTI

...

Link Panes

VectorNTI

...

VectorNTI: Molecule Editing

Close My pBR322

Formatting the graphics of a molecule:

» NCBI nucleotide database: NM_001025366


...

Retrieve directly from

NCBI into VectorNTI:

» Tools » Open » Retrieve...

NM_001025366


...


...Ctrl + click

» ARRANGEMENT


...

Graphics Display Setup

>> all features will be

displayed in their specific way


...

Feature Display Setup

>> change the way this

feature is displayed


...


...Picture

Editing Mode


...


Molecule display setup:

» deselect restriction map to hide the RE sites

Graphics display setup: new style for CDS feature

» Shape 0 + More... (fill color + darker color for border)

Save the molecule in the database


...

Change

graphical

display


...

VectorNTI: Restriction enzymes

...


Import enzymes

» From archive » DBINIT.EA4

» From REBASE » baroich.nnn

Restriction map setup

Restriction analysis and report

Restriction digestion and gel display

Open pUC19 from the database


...

Primer design and PCR analysis

...


...

Select 2 enzymes (use Ctrl)

e.g. PvuI and EcoRV


...


...

Restriction Analysis and Report


...


...

Select BamHI and PvuI


...


Primer design for PCR on GST from plasmid pGEX-5G/LIC

GST: 258-935 bp | MCS: BamHI (951), Xma (956), EcoRI (961)

Molecule construction: GST in pET104-DEST (NdeI – XmaI)


...


With the Fragment Wizard you can construct

a new DNA molecule

Define the needed fragments for the

Goal Molecule Definition list

Molecule construction:

cloning GST in pET104-DEST (NdeI – XmaI)


...


...

hold shift key and click in RE!

else Fragment wizard reverts to 5’!


...


...

hold shift key and click in RE!

else Fragment wizard reverts to 5’!


...


Exercises

» Amplify the SV40 promoter from plasmid pRSVneo

Add a HindIII site to the 5’ end

and a SmaI site to the 3’ end

Save the result as SV40prom

» Clone the SV40 promoter using the

HindIII-SmaI in pUC18

Save as pUC18-SV40

vnti11 basics course

Technology

vectornti explorer subsets

vectornti explorer http

data vectornti

vectornti explorer backup

vectornti explorer summary

vectornti explorer new

main database

subset molecules