Does only one sin in your entire life disqualify you for the kingdom of God?

Because you did not specify denomination, I will present the answer of the Catholic Church.

If you commit venial sin, you will suffer temporal consequences (possibly for the rest of your earthly life) and damage your friendship with God, but you will not end your friendship with God. You can obtain forgiveness through prayer, almsgiving, fasting, and sacramentals.

If you commit mortal sin, God will interpret it as a decision to end your friendship with Him and, absent either perfect contrition or absolution from a priest, a free will decision to enter into a state of eternal disgrace after death. "Many of those who sleep in the dust of the earth shall awake, some to everlasting life, and some to shame and everlasting contempt." (Daniel 12:2 NRSVCE)

Mortal sin is defined as an action that is simultaneously the following three:

1. Grave matter (see Canon law)
2. Done after being informed that it is grave matter
3. Done with full consent of the will

Venial sin is defined as sin that is not mortal.

My answer to this is simple, and while "spiritual", my personal focus is based on how I believe Jesus revealed spiritual Reality to be, rather than on man's religion.

The short answer is "Absolutely Yes ... BUT!". I'll explain:

God's ways are Life (John 14:6, 3:16, 17:3, Deut.30:19, etc). Therefore, following His Ways leads to life.

Not only are God's ways Life, but EVERY good thing (including life) comes only from God (James 1:16-17).

Therefore, to turn away from God's Ways is to turn away from Life -- and turning away from Life means to turn toward death. That is why God said to Adam and Eve, that if they went against His commandment to not eat from the Tree of the Knowledge of Good and Evil, "in the day you eat of it, you will surely die" is how it's frequently translated, but in Hebrew it actually says, "in the day you eat of it, dying, you will die" -- in other words, you will introduce death into yourselves (Gen.2:17)

To sin is to turn away from God, which is to turn away from Life, which is to turn to death. You can go your whole life not taking cyanide, and all you have to do it take it one time, and you are dead. It doesn't matter if you ate organic and exercised your whole life other than that or not, you took death into yourself one time, and you are dead. That's just reality, in the spiritual world as well as the physical world.

God is not looking for a chance to throw lightening bolts at people the first time they screw up. He is not a perfectionist without mercy who gets irritated at any little thing and takes any chance to boot people out of heaven. FAR from it! He went to the most extreme of lengths to keep you and I from eternal death. He started out speaking to mankind directly, warning us of how to avoid death and live forever (Adam and Eve); then when people's hearing began to grow dull, He sent them prophet after prophet to speak His words of warning, and how to gain life; eventually even going so far as to send His sinless Only-begotten Son to die the death in our place, so that we wouldn't all perish but could all of us have the opportunity to live in His glorious Love and Life with Him for all eternity.

On the one hand, we must realize the dire condition we are in because "ALL have sinned and fallen short of the glory of God" (Rom.3:23), and "the wages [automatic result] of sin is death" (Rom.6:23a); and yet on the other hand, all the more the amazingly Good News because of the previous terrible news, God in His good pleasure and everlasting Love "justified (us) freely by His grace through the Redemption that is in Christ Jesus" (Rom 3:24; Luke 12:32).

The basic principle in the Hebrew scriptures is, if one sins, one dies:

Ezekiel 18
⁴ Behold, all souls are mine; … the soul that sinneth, it shall die.

This is repeated in the Greek scriptures, with an escape clause:

Romans 6
²³ For the wages of sin is death, but the free gift of God is eternal life through Christ Jesus our Lord.

The statement "his perfect righteousness is imputed unto you as if you lived a sinless life" isn't exactly true.

Jesus died, even though he never sinned. His immense sacrifice is so great that it can be used to pay the penalty for everyone else.

1 Corinthians 15
³ … Christ died for our sins according to the scriptures

But it isn't a blanket gift to everyone. One must be baptized and accept God's holy spirit, admit one's sins, repent (change one's behaviour and attitude), and then ask for his forgiveness.