The workers are protesting over plans to make them fill in forms accounting for each hour they work. They claim it will lead to excessive monitoring of staff, de-skilling and exposure to repetitive strain injury.
The 24-hour action is expected to be the first of a summer of disruption by the civil servants.
Tony Robertson, Merseyside general secretary of the Public and Commercial Services union (PCS), said: "This is all over the new management system which makes staff account for what they do every hour. It takes a lot for mild-mannered civil servants to go out on strike but this has made everyone incredibly angry and caused a lot of stress and pressure. It has made people who have worked here twenty and thirty years very despondent."
They feel the system is bearing down on them, at a time when the government is forcing through over 100,000 redundancies in the civil service.
"We thought we had an agreement ten days ago but they withdrew their offer. We want them to discuss this with us in a sensible way.
"This is not what we wanted to happen. Many of the workers at Bootle are on a modest wage and losing a day's pay is not something we would agree to lightly. But I fear this could go on throughout the summer."
The PCS said more than 8,000 staff had walked out nationally but the government department insisted the figure was nearer 4,000 and called the action "unwarranted".
A spokesman for Revenue and Customs said: "We are very disappointed with the PCS decision to reinstate industrial action having initially called it off.
"We regard this action as unnecessary and unwarranted and will do all we can to minimise disruption. All significant personnel issues have been successfully negotiated with the union. However, there remain three issues in dispute that would fundamentally affect our ability to manage effectively."
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